CN114008205A - 前导序列 - Google Patents
前导序列 Download PDFInfo
- Publication number
- CN114008205A CN114008205A CN202080042046.3A CN202080042046A CN114008205A CN 114008205 A CN114008205 A CN 114008205A CN 202080042046 A CN202080042046 A CN 202080042046A CN 114008205 A CN114008205 A CN 114008205A
- Authority
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- China
- Prior art keywords
- pvc
- effector
- seq
- payload
- leader sequence
- Prior art date
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- Pending
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Abstract
本发明提供了光杆状菌毒力盒(PVC)效应子前导序列的用途，用于将有效载荷包装到PVC针状复合物中，以及用于制造包装的PVC针状复合物的相关方法。有效载荷是选自多肽、核酸或其组合的一种或多种，并且前导序列和有效载荷形成不同于野生型PVC效应子蛋白的效应子融合体。
Description
本发明涉及前导序列，以及前导序列用于将分子包装成蛋白质复合物的用途。
生物分子(例如肽、蛋白质和核酸)具有作为广泛适用的治疗剂的巨大潜力。实际上，近年来，制药工业存在从“小分子”药物转向更复杂的大分子治疗剂(又名“生物制剂”)的趋势。此类生物制剂包括基于蛋白质的治疗剂(特别是抗体、激素、生长因子和细胞因子)和基于核酸的治疗剂(如短干扰RNA、DNA/RNA疫苗和基因疗法)。
虽然近年来生物制剂市场已经显著发展，但是有效递送系统(以及用于制造此类递送系统的可行方法)的低可用性限制了此类生物治疗剂的分子靶标的多样性，特别是当靶标是胞质时。实际上，市场上大多数批准的肽治疗剂通过靶向胞外组分来起作用，如膜受体或分泌的分子(例如，存在于间隙空间中)。例如，humira(最成功的治疗性单克隆抗体)靶向胞外分泌的细胞因子TNFα。胰岛素通过结合其存在于细胞膜上的同源受体起作用(其他激素肽治疗剂也是相同的情况)。
类似的问题存在于农业工业中，其中基于蛋白质的杀虫剂通常是必须靶向害虫细胞的胞外组分的毒素。举例来说，苏云金芽孢杆菌(Bacillus thuringiensis)毒素是常用的天然杀虫剂，其必须结合膜受体以发挥其毒性作用。
已经开发了胞质递送生物分子的方法用于实验室研究，其通常涉及在将其有效载荷排空到胞质之前，递送与细胞质膜融合的脂质载体内的分子。然而，此类方法在医学和兽医中的用途有限，例如由于它们将分子递送至细胞的非特异性性质。
考虑到细菌分泌系统将分子分泌(或更具体地“注射”)到靶细胞中的天然能力，细菌分泌系统已被研究作为潜在的递送系统。研究最多的此类分泌系统是III型分泌系统(T3SS)，一种在几种革兰氏阴性细菌中发现的“蛋白质附属物”。然而，这些系统的显著缺点是它们始终保持与细菌膜结合，需要使用实际的细菌细胞(包含分泌系统)作为递送系统。因此，难以完全控制什么分子从细菌转移到靶细胞(即使当目标生物制剂过表达时)，因为这些分泌系统通过在细菌的胞质和靶细胞的胞质之间提供连接(例如通道)来起作用，其他组分(可能对宿主有害)可以通过该连接流动。
因此，不仅需要改进的递送系统，而且还需要用于生产此类系统的方法，所述系统发现与具有一系列尺寸和分子性质的分子(有效载荷)是相容的。
本发明解决了上述问题中的一个或多个。
本发明基于以下令人惊讶的发现：光杆状菌属(Photorhabdus)细菌的产毒性光杆状菌毒力盒(PVC)效应子蛋白包含先前未知的“前导序列”(或“前导肽”)，其用于将PVC效应子包装(或“负载”)到所谓的PVC针状复合物(例如“纳米注射器”)中，所述PVC针状复合物随后将PVC效应子递送至靶细胞，在那里它发挥其产毒作用(PVC效应子代表这种纳米注射器的有效载荷)。此外，本发明人已经发现，这样的前导序列实际上可以用于引导与其连接的有效载荷被包装到PVC针状复合物(和相关/同源复合物)中，这是一种充分表征的光杆状菌的分子递送系统。因此，新发现的前导序列令人惊讶地起到以分子有效载荷(或“弹头”)负载PVC针状复合物的作用。
除了这一发现之外，发明人已经开发了这种前导序列用于将“异源”有效载荷(包括非光杆状菌分子)包装/加载到PVC针状复合物中的有利的实际效用，而与异源有效载荷的大小、分子性质或起源无关。
在第一个方面中，本发明提供了光杆状菌毒力盒(PVC)效应子前导序列用于将有效载荷包装到PVC针状复合物中的用途；
其中有效载荷是选自多肽、核酸或其组合(优选多肽)的一种或多种；和
其中前导序列和有效载荷形成不同于野生型PVC效应子蛋白的效应子融合体。
在一个方面中，本发明的一个方面提供了PVC效应子前导序列用于将有效载荷包装到PVC针状复合物中的用途；
其中有效载荷是选自多肽、核酸或其组合(优选多肽)的一种或多种；和
其中前导序列和有效载荷形成不同于PVC效应子蛋白(例如野生型PVC效应子蛋白)的融合体。
换句话说，在一个方面中，本发明提供了一种用于将有效载荷包装到具有PVC效应子前导序列的PVC针状复合物中的方法，该方法包括使(效应子)融合体与PVC针状复合物接触，其中该有效载荷是选自多肽、核酸、或其组合(优选多肽)中的一种或多种；并且其中前导序列和有效载荷形成(效应子)融合体，其不同于PVC效应子蛋白(例如野生型PVC效应子蛋白)。
在由前导序列和有效载荷形成的(效应子)融合体(并且不同于野生型PVC效应子蛋白)的内容中，术语“融合体”和“效应子融合体”在本文中可互换使用。
如实施例中所概述的，通过在细胞(例如宿主细菌细胞)中表达效应子融合体(用检测标记物标记)和PVC针状复合物，分离PVC针状复合物，然后经由检测标记的蛋白质印迹检测来检测PVC针状复合物(例如其破坏形式)内有效载荷的存在或不存在，从而证明了(前导序列的)这种用途，其中效应子融合体被包装到PVC针状复合物中(经由前导序列)中。仅与前导序列融合时，检测到有效载荷的存在，但有效载荷缺乏前导序列时，未检测到有效载荷的存在。
术语“PVC效应子前导序列”意指来自PVC效应子多肽的前导区(多肽区)，其能够将有效载荷(例如效应子)包装到PVC针状复合物中，并且优选是PVC效应子的氨基酸1-50，或省略初始甲硫氨酸时是氨基酸2-50。发明人已经证明了前导序列包含在大量鉴定的PVC效应子多肽序列的氨基酸1-50内(或可以基本上由其组成)。然而，旨在涵盖具有替代长度和定位在PVC效应子内的前导序列(例如，条件是所述前导序列能够将有效载荷包装到PVC针状复合物中)。
PVC效应子的剩余(非前导序列)部分在本文中称为“效应子部分”(例如有效载荷)。效应子部分优选包含PVC效应子蛋白的氨基酸51-C末端或基本上由其组成。
因此，在一个实施方案中，PVC效应子前导序列包含在PVC效应子多肽的氨基酸1-50或2-50(优选1-50)内。
在一个实施方案中，PVC效应子前导序列包含(或基本上由其组成)PVC效应子多肽的氨基酸1-50或2-50(优选1-50)。
术语“野生型PVC效应子蛋白”与术语“内源性PVC效应子蛋白”同义使用，或简称“PVC效应子蛋白”，并且是指具有与效应子部分(例如有效载荷，优选PVC效应子蛋白的氨基酸51-C末端)缔合的内源性前导序列(即对于给定的PVC效应子，优选PVC效应子的氨基酸1-50是内源性的)的(例如完整的)PVC效应子序列。野生型PVC效应子的实例可包含选自SEQID NO.:1-SEQ ID NO.:46的一个或多个序列的氨基酸序列(或基本上由其组成)。因此，本文所述的本发明的融合体/效应子融合体不同于PVC效应子蛋白(例如野生型PVC效应子蛋白)，因为前导序列不与在野生型PVC效应子蛋白的情况下它可以与之融合的效应子部分融合。举例来说，融合体/效应子融合体可以包含与hvnA(基因Plu1649)PVC效应子蛋白的效应子部分(例如SEQ ID NO.:46的氨基酸51-295)融合的“Pnf”PVC效应子蛋白的前导序列(例如SEQ ID NO.:78的前导序列)，但不旨在指与Pnf PVC效应子蛋白的效应子部分(例如SEQID NO.:32的氨基酸51-340)融合的“Pnf”PVC效应子蛋白的前导序列(例如SEQ ID NO.:78的前导序列)。
另一方面，融合体/效应子融合体可以包含与非效应子部分(例如非光杆状菌蛋白，如Ore重组酶)融合的例如“Pnf”PVC效应子蛋白的前导序列(例如SEQ ID NO.:78的前导序列)。因此，前导序列可用于将一系列例如异源(非野生型)物质包装到PVC针状复合物中，打开了使用PVC针状复合物作为模块化、多样化递送系统的可能性，首次用于不仅将天然效应子而且将“非天然”有效载荷递送至细胞。因此，可以制造具有选择的有效载荷的PVC针状复合物。
本发明的另一个方面提供了一种用于制造包括有效载荷的PVC针状复合物的方法(例如，换句话说，一种用于制造包装的PVC针状复合物的方法)，该方法包括：
a.使PVC针状复合物与效应子融合体接触(例如在宿主细胞内)，所述效应子融合体包含与有效载荷融合的PVC效应子前导序列；
b.其中有效载荷是选自多肽、核酸或其组合的一种或多种(优选多肽)；和
c.其中该效应子融合体不同于野生型PVC效应子蛋白。
本发明的一个方面提供了一种用于制造包含有效载荷的PVC针状复合物的方法(例如，换句话说，一种用于制造包装的PVC针状复合物的方法)，该方法包括：
a.使PVC针状复合物与融合体接触(例如在宿主细胞内)，所述融合体包含与有效载荷融合的PVC效应子前导序列，其中前导序列和有效载荷形成不同于PVC效应子蛋白(例如野生型PVC效应子蛋白)的融合体；和
b.其中有效载荷是选自多肽、核酸或其组合的一种或多种(优选多肽)。
在一个实施方案中，所述接触可以在细胞(例如细菌宿主细胞)内、在细胞裂解物中或在纯化的细胞裂解物中(优选在细胞内)发生。在一个实施方案中，所述接触可以在无细胞表达系统内发生。类似地，本文所述的用途可以包括在细胞(例如细菌宿主细胞)内、在细胞裂解物、无细胞表达系统中或在纯化的细胞裂解物中(优选在细胞内，更优选在细菌宿主细胞内)发生的(在融合体/效应子融合体和PVC针状复合物之间的)接触步骤。
编码PVC针状复合物的盒(操纵子)可以与第一启动子可操作地连接，并且编码融合体/效应子融合体(有效载荷)的基因可以与第二(优选不同的)启动子可操作地连接。在一个实施方案中，所述第一和/或第二启动子是诱导型启动子(例如阿拉伯糖诱导型启动子，如pBAD和/或IPTG诱导型启动子)。因此，本发明包括表达系统，其中编码PVC的操纵子存在于第一载体/质粒内(任选地与第一启动子可操作地连接)，并且编码效应子融合体的序列(与有效载荷融合的前导序列)存在于第二(优选不同的)质粒内(任选地与第二启动子连接)。
在一个实施方案中，PVC针状复合物和/或(优选和)效应子融合体可以在选自细菌细胞、酵母细胞、昆虫细胞和/或哺乳动物细胞的一种或多种宿主中表达。在优选实施方案中，PVC针状复合物和效应子融合体可以在选自细菌细胞、酵母细胞、昆虫细胞和哺乳动物细胞(优选细菌细胞)的宿主细胞中一起表达。合适的哺乳动物细胞包括HEK293细胞和/或CHO细胞。
PVC针状复合物和/或(优选和)效应子融合体(有效载荷)可以在异源细菌表达系统(优选大肠杆菌)中表达。在一个实施方案中，PVC针状复合物和/或(优选和)PVC效应子可以在光杆状菌细胞中表达，任选其中光杆状菌细胞的PVC操纵子对于细胞是内源性的(并且任选其中PVC操纵子可操作地连接诱导型启动子，所述诱导型启动子可以整合到基因组中以通过基因工程可操作地连接PVC操纵子)。例如，可以将诱导型启动子引入光杆状菌细胞的基因组中PVC(操纵子)的5'端，优选通过如实施例(例如实施例3)中所述的重组工程来进行。
有效载荷可以是例如治疗性有效载荷，使得PVC针状复合物可用于医学治疗。
在再一个方面中，本发明提供了一种(包装的)PVC针状复合物，其用于治疗方法中；
a.其中PVC针状复合物包含效应子融合体(例如与其一起包装)，所述效应子融合体包含(或基本上由其组成)与有效载荷融合的PVC效应子前导序列；
b.其中所述有效载荷是选自多肽、核酸或其组合的一种或多种(优选多肽)；和
c.其中该效应子融合体不同于野生型PVC效应子蛋白。
本发明的另一个方面提供了一种(包装的)PVC针状复合物，其用于治疗方法中；
a.其中PVC针状复合物容纳融合体(例如与其一起包装)，所述融合体包含(或基本上由其组成)与有效载荷融合的PVC效应子前导序列；
b.其中所述有效载荷是选自多肽、核酸或其组合的一种或多种(优选多肽)；和
c.其中该融合体不同于PVC效应子蛋白(例如野生型PVC效应子蛋白)。
在一个方面中，本发明提供了一种治疗受试者的方法，该方法包括向受试者(例如患者)施用(包装的)PVC针状复合物；
a.其中PVC针状复合物包含效应子融合体(例如与其一起包装)，所述效应子融合体包含(或基本上由其组成)与有效载荷融合的PVC效应子前导序列；
b.其中所述有效载荷是选自多肽、核酸或其组合(优选多肽)的一种或多种；和
c.其中该效应子融合体不同于野生型PVC效应子蛋白。
换句话说，本发明的一个方面提供了一种治疗受试者的方法，该方法包括向受试者(例如患者)施用(包装的)PVC针状复合物；
a.其中PVC针状复合物容纳融合体(例如与其一起包装)，所述融合体包含(或基本上由其组成)与有效载荷融合的PVC效应子前导序列；
b.其中所述有效载荷是选自多肽、核酸或其组合的一种或多种(优选多肽)；和
c.其中该融合体不同于PVC效应子蛋白(例如野生型PVC效应子蛋白)。
在优选实施方案中，有效载荷是多肽。
受试者可以是哺乳动物受试者，优选人受试者。
术语“PVC针状复合物容纳效应子融合体”和“PVC针状复合物包含效应子融合体”是指具有包装的效应子融合体的PVC针状复合物，或者换句话说，与效应子融合体一起包装的PVC针状复合物。
术语“包装的效应子融合体”、“融合体”和“效应子融合体”(例如其中融合体/效应子融合体不同于野生型PVC效应子蛋白)包括PVC效应前导序列和包装到PVC针状复合物中后保持接触(例如融合)的有效载荷的组合(例如前导序列尚未从有效载荷切割出来)，以及PVC效应前导序列和不再直接接触(例如不再融合，如前导序列从有效载荷切割后)的有效载荷的组合。
如本文所用的术语“治疗(treat)”或“治疗(treating)”涵盖预防性治疗(例如预防疾病发作)以及矫正性治疗(治疗已经患有疾病的受试者)。优选，如本文所用的“治疗(treat)”或“治疗(treating)”意指矫正性治疗。术语“治疗(treat)”或“治疗(treating)”涵盖治疗疾病及其症状两者。在一些实施方案中，“治疗(treat)”或“治疗(treating)”是指疾病的症状。
因此，PVC针状复合物可以以治疗有效量或预防有效量施用于受试者。
“治疗有效量”是(包装/负载的)PVC针状复合物的任何量，其在单独或组合(例如，与另一种治疗剂，并行或连续施用并且叠加或协同作用)施用于受试者以治疗疾病(或其症状)时足以实现疾病或其症状的这种治疗。
“预防有效量”是(包装/负载的)PVC针状复合物的任何量，其在单独或组合(例如，与另一种治疗剂，并行或连续施用并且叠加或协同作用)施用于受试者时，抑制或延迟疾病(或其症状)的发作或复发。在一些实施方案中，预防有效量完全预防疾病的发作或复发。“抑制”发作意指降低疾病发作(或其症状)的可能性，或完全预防发作。
在相关方面中，提供了一种(包装的)PVC针状复合物，其包含(例如，容纳/与其一起包装)效应子融合体；
a.其中所述效应子融合体包含(或基本上由其组成)与有效载荷融合的PVC效应前导序列(或换句话说，其中所述效应子融合体由PVC效应前导序列和有效载荷形成)；
b.其中所述有效载荷是选自多肽、核酸或其组合的一种或多种；和
c.其中该效应子融合体不同于野生型PVC效应子蛋白。
换句话说，本发明的一个方面提供了一种(包装的)PVC针状复合物，其容纳(例如与其一起包装)融合体；
a.其中所述融合体包含(或基本上由其组成)与有效载荷融合的PVC效应子前导序列(或换句话说，其中所述融合体由PVC效应子前导序列和有效载荷形成)；
b.其中所述有效载荷是选自多肽、核酸或其组合的一种或多种(优选多肽)；和
c.其中该融合体不同于PVC效应子蛋白(例如野生型PVC效应子蛋白)。
在优选实施方案中，(包装的)PVC针状复合物是分离的(例如非天然的)PVC针状复合物。
如下所述，PVC针状复合物通常在自然界中起作用以将产毒的PVC效应子递送至昆虫靶标。通过极大地扩展可以包装到PVC针状复合物中的有效载荷的数量和种类，本发明同时扩展了可以被靶向和杀灭的无脊椎动物(例如害虫)的数量和种类，如变形虫、线虫、蠕虫和昆虫。
在本发明的再一个方面中，提供了一种用于抑制害虫的方法，该方法包括：
a.使害虫或包含害虫的目标区域与包含(例如容纳/包装有)效应子融合体的(包装的)PVC针状复合物接触；
b.其中效应子融合体包含(或基本上由其组成)与有效载荷融合的PVC效应子前导序列(或换句话说，其中所述效应子融合体由PVC效应子前导序列和有效载荷形成)；
c.其中所述有效载荷是选自多肽、核酸或其组合的一种或多种(优选多肽)；和
d.其中该效应子融合体不同于野生型PVC效应子蛋白。
本发明的一个方面提供了一种用于抑制害虫的方法，该方法包括：
a.使害虫或包含害虫的目标区域与容纳(例如包装有)融合体的(包装的)PVC针状复合物接触；
b.其中融合体包含(或基本上由其组成)与有效载荷融合的PVC效应子前导序列(或换句话说，其中所述融合体由PVC效应子前导序列和有效载荷形成)；
c.其中所述有效载荷是选自多肽、核酸或其组合的一种或多种(优选多肽)；和
d.其中该融合体不同于PVC效应子蛋白(例如野生型PVC效应子蛋白)。
术语“PVC针状复合物容纳效应子融合体”和“包含效应子融合体的PVC针状复合物”意指具有包装的效应子融合体的PVC针状复合物。
术语“目标区域”是指存在害虫和/或可能(例如，预期或怀疑)存在害虫的区域。
因此，在一个实施方案中，可以在存在害虫之前和/或存在害虫时接触目标区域。目标区域可以在害虫附近(例如，非常接近)。可替代地，目标区域可以是用户希望保护免受害虫影响的区域。例如，目标区域可以包括植物和/或植物产品。
术语“遏制害虫”包括“害虫防治”、“抑制害虫生长”、“抑制害虫增殖”和/或“害虫死亡”。
此类害虫的实例包括一种或多种昆虫、螨虫、猪蝽、大蝽、蜈蚣、软体动物、千足虫、原生生物、真菌、蠕虫和/或血源性寄生虫。害虫可以处于任何发育阶段，例如可以是幼虫和/或成年害虫(例如成虫)。
本发明可用于靶向各种农业、商业、家庭和花园害虫。
在一个实施方案中，害虫是昆虫、螨虫、猪蝽、大蝽、蜈蚣、软体动物和/或千足虫。合适地，害虫可以是昆虫和/或螨虫(优选昆虫)。
合适的昆虫的实例包括鳞翅目(Lepidoptera)、鞘翅目(Coleoptera)、双翅目(Diptera)、蜚蠊目(Blattodea)、膜翅目(Hymenoptera)、等翅目(Isoptera)、直翅目(Orthoptera)、缨尾目(Thysanura)和/或革翅目(Dermaptera)的昆虫。在一个实施方案中，鳞翅目的昆虫可以是蛾和/或蝴蝶中的一种或多种。合适的蛾包括烟草天蛾(ManducaSexta)和/或大蜡螟(Galleria mellonella)。
在一个实施方案中，鞘翅目昆虫可以是欧洲金龟子蛴螬、北方伪装金龟子蛴螬、南方伪装金龟子蛴螬、日本甲虫蛴螬、六月甲虫蛴螬、黑藤象鼻虫、草莓根象鼻虫、粘土色象鼻虫、科罗拉多马铃薯甲虫和/或金针虫中的一种或多种。在另一个实施方案中，双翅目昆虫可以是长脚蝇的蛆(例如大蚊的幼虫)、洋葱蛆、卷心菜蛆、胡萝卜锈病蝇蛆、真菌幼虫和/或蚊子中的一种或多种。在另一个实施方案中，蜚蠊目昆虫可以是蟑螂，合适地选自美洲蟑螂和/或德国蟑螂的一种或多种蟑螂。
在一个实施方案中，膜翅目昆虫可以是蚂蚁。合适地，蚂蚁可以是木蚁、有气味的家蚁、铺道蚁、阿根廷蚁、法老蚁、黄褐色疯狂蚁、收获蚁、进口红火蚁、南方火蚁、欧洲火蚁和/或小火蚁中的一种或多种。在另一个实施方案中，膜翅目昆虫可以是黄蜂。
在一个实施方案中，等翅目昆虫可以是白蚁。合适地，白蚁可以是湿木白蚁、干木白蚁和/或地下白蚁中的一种或多种。在另一个实施方案中，直翅目昆虫可以是蟋蟀、蝗虫和/或蝗虫中的一种或多种。在一个实施方案中，缨尾目昆虫可以是衣鱼。在另一个实施方案中，革翅目昆虫可以是蠼螋。
合适的软体动物的实例包括蛞蝓和/或蜗牛。
在一个实施方案中，害虫是原生生物。在一个实施方案中，所述原生生物是选自巨形变形虫(Chaos carolinense)、变形阿米巴(Amoeba proteus)、福氏内格里阿米巴原虫(Naegleria fowleri)、盘基网柄菌(Dictyostelium discoideum)、溶组织内阿米巴(Entamoeba histolytica)、阴道毛滴虫(Trichomonas vaginalis)、人芽囊原虫(Blastocystis hominis)、利什曼原虫属(Leishmania Spp.)和兰伯贾第鞭毛虫Giardialamblia中的一种或多种。在一个实施方案中，所述原生生物是Fonticula alba、盘基网柄菌(Dictyostelium discoideum)、莱茵衣藻(Chlamydomonas reinhardtii)、Crytomonasparamedium、载色宝琳虫(Paulinella chromatophora)、Nannochloropsis gaditana和/或四膜虫属(Tetrahymena Spp.)的一种或多种。
在一个实施方案中，害虫是真菌。在一个实施方案中，所述真菌是选自兔脑炎原虫(Encephalitozoan cuniculi)、Nasema apis、东方蜜蜂微孢子虫(Namema ceranae)、Vittaforma carneae、Enterocytosoan bieneusi、Spraguea lophii、Vavra culiculis、Edharzardia aedes、Nematocida parisii、Razella Spp.、Parasitella parasitica、Lichteimia ramose、Sporisorium scitamineum、变色栓菌(Trametes versicolor)和/或粗环点革菌(Punctularia strigosozonata)的一种或多种真菌。
在一个实施方案中，所述真菌是假丝酵母属(Candida spp.)。所述假丝酵母属可以是选自白色假丝酵母(C.albicans)、阿斯卡拉蚜假丝酵母(C.ascalaphidarum)、两栖假丝酵母(C.amphixiae)、南极假丝酵母(C.Antarctica)、阿根廷假丝酵母(C.argentea)、大西洋假丝酵母(C.atlantica)、大气假丝酵母(C.atmosphaerica)、耳假丝酵母(C.auris)、钝顶假丝酵母(C.blattae)、凤梨假丝酵母(C.bromeliacearum)、嗜果假丝酵母(C.carpophila)、香芹假丝酵母(C.carvajaiis)、天牛假丝酵母(C.cerambycidarum)、查氏假丝酵母(C.chauliodes)、延胡索假丝酵母(C.corydalis)、多西假丝酵母(C.dosseyi)、都柏林假丝酵母(C.dubliniensis)、C.ergatensis、果实假丝酵母(C.fructus)、光滑假丝酵母(C.glabrata)、发酵假丝酵母(C.fermentati)、吉利蒙假丝酵母(C.guilliermondii)、希木龙假丝酵母(C.haemulonii)、扁平云假丝酵母(C.humilis)、昆虫门假丝酵母(C.insectamens)、昆虫假丝酵母(C.insectorum)、中型假丝酵母(C.intermedia)、杰弗里希假丝酵母(C.jeffresii)、乳酒假丝酵母(C.kefyr)、煤油假丝酵母(C.keroseneae)、克鲁斯氏假丝酵母(C.krusei)、葡萄牙假丝酵母(C.lusitaniae)、C.lyxosophila、麦芽糖假丝酵母(C.maltose)、海生假丝酵母(C.marina)、膜醭假丝酵母(C.membranifaciens)、蒙奇假丝酵母(C.mogii)、橄榄假丝酵母(C.oleophila)、俄勒冈假丝酵母(C.oregonensis)、近平滑假丝酵母(C.parapsilosis)、橡树假丝酵母(C.quercitrusa)、柱状假丝酵母(C.rugose)、清酒假丝酵母(C.sake)、休哈塔假丝酵母(C.shehatea)、C.temnochilae、纤维假丝酵母(C.tenuis)、C.theae、C.tolerans、C.tropicalis(热带假丝酵母)、C.tsuchiyae、C.sinolaborantium、大豆假丝酵母(C.sojae)、C.subhashii、维斯假丝酵母(C.viswanathii)、产朊假丝酵母(C.utilis)、C.ubatubensis和/或泽普林假丝酵母(C.zemplinina)的一种或多种。合适地，所述假丝酵母属可以是白色假丝酵母。
在另一个实施方案中，害虫是蠕虫。所述蠕虫可以是选自环节动物门(Annelida)、扁形动物门(Platyhelminthes)、线虫门(Nematoda)和/或棘头动物门(Acanthocephala)的一种或多种。在一个实施方案中，所述蠕虫是寄生性扁虫。所述寄生性扁虫可以是选自绦虫属(Cestoda)、吸虫属(Trematoda)和/或单殖吸虫属(Monogenea)的一种或多种。在一个实施方案中，所述蠕虫是寄生性线虫。所述寄生性线虫可以是选自蛔虫(蛔虫属(Ascaris))、丝虫、钩虫、蛲虫(蛲虫属(Enterobius))和/或鞭虫(毛首鞭虫(Trichuris trichiura))的一种或多种。
在一个实施方案中，害虫是血源性寄生虫。所述血源性寄生虫可以是选自锥虫属(Trypanosoma Spp)(例如布氏锥虫(Trypanosoma brucei)和/或克氏锥虫(T.cruzi))、巴贝虫属(Babesia Spp)(例如田鼠巴贝虫(Babesia microti))、利什曼原虫属(LeishmaniaSpp)、疟原虫属(Plasmodium Spp)(例如恶性疟原虫(P.falciparum))和/或弓形虫属(Toxoplasma Spp)(例如刚地弓形虫(Toxoplasma gondii)的一种或多种。
用于害虫防治的PVC针状复合体合适地是环境安全的(例如环境安全的杀虫组合物)。
其他有利的效用包括例如在实验室研究期间将有效载荷递送至细胞。这样的细胞可以是体外细胞系的一部分，或者可以是动物(例如研究动物模型)的细胞。另外地或可替代地，细胞可以包含在离体系统(例如类器官)内。
本发明的另一个方面提供了用于将有效载荷递送到细胞中的体外(和/或离体)方法，所述方法包括：
a.使细胞与(包装的)PVC针状复合物接触，所述PVC针状复合物包含(例如容难/包装有)效应子融合体；
b.其中效应子融合体包含(或基本上由其组成)与有效载荷融合的PVC效应子前导序列(或换句话说，其中所述效应子融合体由PVC效应子前导序列和有效载荷形成)；
c.其中所述有效载荷是选自多肽、核酸或其组合的一种或多种(优选多肽)；和
d.其中该效应子融合体不同于野生型PVC效应子蛋白。
本发明的一个方面提供了用于将有效载荷递送到细胞中的体外(和/或离体)方法，该方法包括：
a.使细胞与容纳(例如包装有)融合体的(包装的)PVC针状复合物接触；
b.其中融合体包含(或基本上由其组成)与有效载荷融合的PVC效应子前导序列(或换句话说，其中所述融合体由PVC效应子前导序列和有效载荷形成)；
c.其中所述有效载荷是选自多肽、核酸或其组合的一种或多种(优选多肽)；和
d.其中融合体不同于PVC效应子蛋白(例如野生型PVC效应子蛋白)。
在一个方面中，本发明提供了效应子融合体，其包含(或基本上由其组成)与有效载荷融合的PVC效应子前导序列(或换句话说，由PVC效应子前导序列和有效载荷形成的效应子融合体)；
a.其中所述有效载荷是选自多肽、核酸或其组合的一种或多种；和
b.其中该效应子融合体不同于野生型PVC效应子蛋白。
本发明的一个方面提供了融合体，其包含(或基本上由其组成)与有效载荷融合的PVC效应子前导序列(或换句话说，由PVC效应子前导序列和有效载荷形成的融合体)；
a.其中所述有效载荷是选自多肽、核酸或其组合的一种或多种(优选多肽)；和
b.其中该融合体不同于PVC效应子蛋白(例如野生型PVC效应子蛋白)。
在一个实施方案中，融合体/效应子融合体是分离的融合体/效应子融合体(例如分离的非天然存在的融合体/效应子融合体)。
本发明包括包含编码融合体/效应子融合体的核苷酸序列的核酸，和/或包含所述核酸的表达载体。还包括包含所述核酸和/或表达载体的宿主细胞。
如上所述，本发明人首次发现并实际利用了本文所述的前导序列。
因此，本发明的另一个方面提供了分离的PVC效应子前导序列(例如，其中分离的PVC效应子前导序列能够将有效载荷包装到PVC针状复合物中)。
在相关方面中，提供了分离的核酸，其包含编码PVC效应子前导序列的核苷酸序列。
分离的PVC效应子前导序列可以是重组的、合成的和/或纯化的。编码PVC效应子前导序列的分离的核酸可以是重组的、合成的和/或纯化的。
下面提供了关于本发明背景和本文使用的术语的更多详细内容。
光杆状菌是肠杆菌属(Enterobacteriace)的细菌，由三种正式公认的(迄今)物种为代表-即发光光杆状菌(P.luminescens)、不对称光杆状菌(P.asymbiotica)和温度光杆状菌(P.temperata)。重要的菌株包括不对称光杆状菌南极(australis)亚种和发光光杆状菌laumondii亚种。目前可用的基因组序列可在GenBank上获得(不对称光杆状菌ATCC43949完整基因组-GenBank登录号:FM162591.1；Photorhabdus laumondii laumondii亚种菌株TT01染色体，完整基因组-GenBank登录号:CP024901.1)。
提及“发光光杆状菌laumondii亚种”，在本文可与“发光光杆状菌laumondii亚种TT01”、“Photorhabdus laumondii laumondii亚种菌株TT01”和“发光杆菌TT011”互换使用。
Wilkinson等描述了另一种不对称光杆状菌菌株，即不对称光杆状菌Kingscliff的基因组序列(FEMS Microbiology Letters，Volume 309，Issue 2，2010年8月，第136-143页)，其通过引用并入本文中。更多的基因组序列描述于Thanwisai等(PLoS ONE 7(9):e43835)，其通过引用并入本文中。
这些物种中的每一种包含至少一个称为光杆状菌毒力盒(PVC)操纵子的操纵子，其编码PVC针状复合物，其在本文中可称为“纳米注射器”。鉴于光杆状菌通常在自然界中从(共生)昆虫病原性异小杆线虫属物种(Heterorhabditis sp.)线虫回流后作为杀昆虫细菌被发现(例如为了避免与昆虫竞争食物和资源)，可以理解PVC针状复合物在自然界中起到抑制昆虫的作用。实际上，已经显示出分离的PVC针状复合物(容纳/包装有天然效应子毒素，例如Pnf)可用于杀死昆虫幼虫-参见实施例2。光杆状菌毒力盒代表至少四种充分表征的光杆状菌毒素递送系统中的一种。光杆状菌蛋白杀虫毒素的其他主要类别包括“毒素复合物”(Tcs)、“二元PirAB毒素”和“使毛虫松软”(Mcf)毒素。
术语“光杆状菌毒力盒”(PVC)(在本文中与术语“PVC操纵子”同义使用)意指光杆状菌基因组的离散操纵子，所述基因组包含编码多肽亚基的基因，所述多肽亚基在表达时组装以提供大分子PVC针状复合物。这些盒的分子结构已经得到充分表征和描述，例如在The Molecular Biology of Photorhabdus Bacteria(Springer InternationalPublishing AG 2017，ISBN:978-3-319-52714-7，第10章，第159-177页)中，通过引用并入本文中。PVC(操纵子)通常包含约16个编码结构蛋白的基因(pvd-pvc16)，所述结构蛋白组装以提供“PVC针状复合物”，其通常随后是在3'末端的一个或多个基因，其编码具有毒性活性的PVC效应子基因(并且通常是典型的T3SS样效应子的同源物)。光杆状菌基因组通常包含多个这样的盒(例如至少四个)，其通常与不同的效应子有效载荷或甚至多个效应子有效载荷相关联。
已经在光杆状菌属和其他属的成员的基因组中观察到三类PVC结构操纵子(I、II和III类)。每个类别内的PVC在编码它们含有的结构蛋白的基因的数量和类型方面是相似的(参见图1(B))。更详细地，I类PVC(其在本文中可称为“原型PVC”)包含16个保守基因(PVC1-16)。II类缺少pvc13宿主细胞结合纤维和pvc3，(不希望受理论束缚)发明人认为其可能是将pvc13纤维蛋白附着到PVC针状复合物(纳米注射器)上的次要特化鞘亚基。因此，认为该类别可以是“非特异性的”，将有效载荷注射到多种(潜在的任何)细胞类型中。III类类似于I类，但在(未知功能)操纵子的起始处具有另外的Pvc0基因，以及在pvc13和pvc14之间编码的两个另外的基因，其类似于“侵袭”型蛋白基因。该类别通常见于光杆状菌属的人临床分离菌株中-发明人已经表明，菌株(携带编码PVC III类操纵子的PVC操纵子)在37℃下生长并暴露于人血清时，可能发生PVC III类的最佳转录，表明该类别可能是PVC针状复合物的哺乳动物适应形式。
图1(D)中显示了示例性盒(PVC)，其显示了不对称光杆状菌ATCC43949(可从ATCC获得，登录号：ATCC 43949)的模型“I类”PVC操纵子的图谱，所述操纵子与下游效应子基因“PAU_03332”(编码Pnf蛋白效应子，例如SEQ ID NO.:32)相关。这种模型操纵子称为PaATcc43949PVCpnf。这个操纵子包含十六个个结构基因(pvc1-16)和两个编码效应子的基因(3'末端)(在这种情况下，为pvc17/Rhs样，编码Rhs样效应子，和pvc21，编码Pnf效应子)。所述基因pvc1-16对应于GenBank登录号FM 162591.1的序列的基因PAU_03353至PAU_03338，并且由SEQ ID NO.:93的序列来表示。
实例PVC操纵子(例如编码结构基因，但不编码PVC效应子)在SEQ ID NO:93(其编码图1(D)中示意性地显示的操纵子)中提供，其他实例是SEQ ID NO:94和SEQ ID NO:95。这些序列在PVC盒/操纵子的第一个结构基因(pvc1)的ATG起始密码子处开始，并且在最终结构基因(pvc16)的TAA终止密码子处结束。
来自类别I-III中任一类别的PVC针状复合物可用于各种应用。然而，某种类别的PVC针状复合物可能特别适合于递送至限定的细胞类型。例如，用于将有效载荷递送至哺乳动物细胞的PVC针状复合物可以合适地是III类成员。用于将有效载荷递送至昆虫细胞(例如递送至昆虫)的PVC针状复合物可以合适地是I类成员(例如由SEQ ID NO.:93编码的不对称光杆状菌PVCpnf，例如在大肠杆菌中从粘粒克隆表达)。
因此，如本领域技术人员将理解的，术语“PVC针状复合物”(在本文中与术语“PVC针状复合物递送系统”和“纳米注射器”同义使用)意指包含由光杆状菌属细菌的PVC(操纵子)编码的多肽亚基的大分子蛋白复合物。PVC针状复合物以纳米注射器结构组装，具有与抗菌R型绿脓菌素类似的物理结构(表面上)(参见图3)。功能和分子研究表明，PVC针状复合物包装有(载有)一种或多种PVC效应子蛋白(即，PVC效应子蛋白被包装在其中或其上)，包装的PVC针状复合物从细菌释放，然后将PVC效应子注射到靶细胞中，使得PVC效应子蛋白可以发挥毒性。
术语“PVC针状复合物”优选涵盖由操纵子编码的PVC针状复合物样结构/复合物，所述操纵子包含与光杆状菌PVC操纵子的基因同源的基因。PVC样元件不限于光杆状菌，并且在昆虫病原细菌嗜虫沙雷氏菌(Serratia entomophila)的pADAP质粒上存在充分表征的同源操纵子(PVC操纵子)。此外，细菌藤黄假单胞菌(Pseudoaiteromonas luteoviolacea)采用类似的和(至少部分)同源的PVC样“注射体”针状复合物系统(例如用于控制海洋蠕虫秀丽水蚤(Hydroides elegans)的变态)。结构存在于其他肠杆菌科(如耶尔森氏菌属(Yersinia Spp.))中，其由与PVC操纵子具有同源性的操纵子编码，并且可以与本文所述的前导序列一起使用。这些(PVC样)结构中的每一种都包含在本文所用的术语“PVC针状复合物”中。
因此，PVC针状复合物是“纳米注射器”复合物，其中由效应子基因编码的多肽被包装(负载)在PVC针状复合物内或在PVC针状复合物的末端(尖端)处，因此代表PVC针状复合物的“有效载荷”或“弹头”。本发明人已经证明，在与靶细胞的膜相互作用并将有效载荷注射到细胞胞质中前，PVC针状复合物本身(具有仍然负载的有效载荷)从光杆状菌细胞自由释放(例如分泌)。实际上，在分离/纯化PVC针状复合物并使用它们抑制(例如杀灭)昆虫幼虫之前，发明人已经成功地在异源表达系统中表达和负载PVC针状复合物(参见实施例2)。因此，PVC针状复合物充当长程蛋白质递送系统。
在一个实施方案中，PVC针状复合物由与选自SEQ ID NO.:93、SEQ ID NO.:94和SEQ ID NO.:95的序列(例如，SEQ ID NO.:93)具有至少75％序列同一性(优选至少85％序列同一性；更优选至少95％序列同一性)的序列编码。
在一个实施方案中，PVC针状复合物由选自SEQ ID NO.:93、SEQ ID NO.:94和SEQID NO.:95的序列(例如，SEQ ID NO.:93)编码。
前导序列/信号序列通常是肽，通常为10-30个氨基酸长，存在于大多数(新)表达的蛋白质的N-末端，其注定走向分泌途径(例如，用于将所述蛋白质引导至细胞膜上的蛋白质传导通道)。许多蛋白质需要进入高尔基体或内质网的信号序列。
在本文的“PVC效应子前导序列”的内容中使用的术语“前导序列”(在本文中与术语“前导肽”、“信号序列”、“靶向信号”、“定位信号”、“定位序列”和“转运肽”可互换使用)意指用于将PVC效应子指引到PVC针状复合物的内部或末端(尖端)中的多肽序列-因此，前导序列用于将PVC效应子包装到PVC针状复合物中。PVC针状复合物随后可以将PVC效应子递送(例如注射)到靶细胞中。PVC针状复合物可以是组装的PVC针状复合物。术语“PVC针状物复合物”可以指PVC针状物复合物的片段(例如，其中前导序列接触所述片段，并且任选地PVC针状物复合物围绕前导序列-有效载荷“效应子融合体”组装)。
PVC前导序列通常存在于PVC效应子或其同系物的N-末端(特征在于头50个氨基酸或包含在头50个氨基酸内)。然而，本发明包括PVC效应子和PVC效应子同源物的前导序列，其可以在除此类PVC效应子/同源物的N-末端区域之外的区域中(例如在C-末端区域中)被发现。
在一个实施方案中，前导序列包含(或基本上由其组成)PVC效应子(例如PVC效应子蛋白)的氨基酸残基1-50。提及“氨基酸残基1-50”包括“氨基酸残基2-50”，其中N-末端甲硫氨酸被省略，例如已被切割。前导序列可以是PVC效应子的N-末端50个氨基酸的片段(例如，包含≤45、≤35、≤25或≤15个氨基酸的片段或基本上由其组成的片段)，条件是该片段能够将有效载荷包装到PVC针状复合物中。
在一个实施方案中，本发明的前导序列(例如，分离的前导序列)包含(或基本上由其组成)与选自SEQ ID NO.:47-SEQ ID NO.:92的一个或多个序列(优选SEQ ID NO.:50、SEQ ID NO.:68、SEQ ID NO.:71、SEQ ID NO.:76、SEQ ID NO.:78或SEQ ID NO.:92)具有至少50％、55％、60％、65％、70％、75％、80％、85％、90％、95％或100％序列同一性的氨基酸序列-例如条件是前导序列能够将有效载荷包装到PVC针状复合物中。在一个优选的实施方案中，前导序列包含(或基本上由其组成)与选自SEQ ID NO.:47-SEQ ID NO.:92的一个或多个序列(优选SEQ ID NO.:50、SEQ ID NO.:68、SEQ ID NO.:71、SEQ ID NO.:76、SEQ IDNO.:78或SEQ ID NO.:92)具有至少60％序列同一性的氨基酸序列-例如条件是前导序列能够将有效载荷包装到PVC针状复合物中。在更优选的实施方案中，前导序列包含(或基本上由其组成)选自SEQ ID NO.:47-SEQ ID NO.:92的一个或多个的氨基酸序列(优选SEQ IDNO.:50、SEQ ID NO.:68、SEQ ID NO.:71、SEQ ID NO.:76、SEQ ID NO.:78或SEQ ID NO.:92)。在一个实施方案中，前导序列包含(或基本上由其组成)选自SEQ ID NO.:47-SEQ IDNO.:92的氨基酸序列(优选SEQ ID NO.:50、SEQ ID NO.:68、SEQ ID NO.:71、SEQ ID NO.:76、SEQ ID NO.:78或SEQ ID NO.:92)。
在一个实施方案中，前导序列包含(或基本上由其组成)选自SEQ ID NO.:50、SEQID NO.:68、SEQ ID NO.:71、SEQ ID NO.:76、SEQ ID NO.:78和SEQ ID NO.:92的氨基酸序列。
在一个实施方案中，前导序列包含SEQ ID NO.:50的氨基酸序列(或基本上由其组成)。在一个实施方案中，前导序列包含SEQ ID NO.:68的氨基酸序列(或基本上由其组成)。在一个实施方案中，前导序列包含SEQ ID NO.:71的氨基酸序列(或基本上由其组成)。在一个实施方案中，前导序列包含SEQ ID NO.:76的氨基酸序列(或基本上由其组成)。在一个实施方案中，前导序列包含SEQ ID NO.:78的氨基酸序列(或基本上由其组成)。在一个实施方案中，前导序列包含SEQ ID NO.:92的氨基酸序列(或基本上由其组成)。
不希望受理论束缚，认为前导序列基于氨基酸性质共享“化学组成共有序列”。更具体地，前导序列包含相似的电荷模式，该模式包含2×个带负电荷的区域，每个之后是带正电荷的区域(例如[-ve][+ve][-ve][+ve])-参见图9。这与2型分泌系统的毒素的前导序列一致，其包含[+ve][疏水性][+ve][C]的电荷/性质模式。进一步的理论认为前导序列共享典型的“螺旋-转角-螺旋”结构。另一种理论是前导序列形成由存在于PVC针状复合物内部或末端(例如尖端)的ATP酶(例如由图1(D)的模型操纵子中的基因PAU_03339(pvc15)编码)识别的结构。
术语“PVC效应子”(与术语“PVC操纵子编码的效应子”和“PVC效应子蛋白”同义使用)意指由光杆状菌PVC操纵子编码的效应子多肽，更特别地(并且通常)在所述操纵子的结构基因的下游(3')不远处被发现(优选地在pvc16的下游的不远处或就在pvc16的下游，并且通常在5kb内)。术语“PVC效应子”优选包括其同系物。因此，前导序列也可以来自由编码PVC效应子的基因的同源物的基因编码的多肽-这类同源物的实例参见表1。实际上，通过检测pvc16下游基因与已知毒素多肽(例如编码所述毒素多肽的基因)的同源性来辅助PVC效应子的鉴定。如本领域技术人员将理解的，术语“同源物”优选意指源自相同祖先基因并且共享相似功能的基因-这样的基因(或由其编码的多肽)与编码PVC效应子的基因同源。同源物可以来自光杆状菌属物种的基因组或来自光杆状菌属物种以外的物种。合适的同源物的实例概述于表1中。
本发明人已经详细阐明和表征了在光杆状菌属的三种最常见(最佳表征的)菌株以及不对称光杆状菌泰国菌株PB68.1中编码这些PVC针状复合物的PVC效应子的基因。这是基于分析遗传连锁与操纵子的PVC结构基因的3'端的接近性以及效应子(例如已知效应子/毒素蛋白的同源物)的蛋白质序列的预测功能来进行的。更详细地，PVC效应子(例如编码PVC效应子的基因)通常被鉴定为与编码已知毒素多肽的基因(例如表1中概述的同源物)具有同源性的开放阅读框(ORF)，并且通常存在于PVC操纵子(例如pvc16)的最终结构基因下游1千碱基至5千碱基(kb)的距离内(例如1kb内)(通常具有很少或没有间插基因)。通常，在操纵子末端(编码PVC针状复合物)和PVC效应子基因之间没有“非毒素样”ORF。尽管在这些区域中可能存在(例如一个或两个)其他小的预测基因，但这些其他基因不被指定为PVC效应子(由于与已知的效应子/毒素基因缺乏同源性，如上所述)。
为了将推定的PVC效应子基因(例如在5kb的距离内的ORF，例如在PVC操纵子的最终结构基因下游1kb内)指定为PVC效应基因，发明人使用了BlastP和HHPRED(https://toolkit.tuebingen.mpg.de/#/tools/HHPRED)的组合。基于与已知毒素编码基因的直接同源性、与毒素蛋白家族的相似性、与PVC操纵子的接近性(例如在PVC操纵子pvc16的最终结构基因下游1-5kb内)和/或基于预测的二级结构与已知毒素的二级结构的结构域相似性，将推定的PVC效应子基因指定为PVC效应子基因。
因此，可以通过(i)鉴定pvc16(例如通过与已知pvc16的序列同源性)，(ii)鉴定pvc16的3'ORF，优选pvc16下游≤5kb)，和(iii)通过鉴定与编码毒素多肽(例如，表1中标记为“同源物”的列中描述的毒素蛋白)的已知基因的序列同源性来确认所述ORF编码PVC效应子来鉴定(在光杆状菌属基因组内的)PVC效应子(基因)。
举例来说，PVC效应子基因PAU_03337(由于与毒力sep基因同源，在本文中称为“sepC”)位于PVC操纵子的pvc16(PAU_03338)下游的325个碱基对(bp)处，所述PVC操纵子在本文中称为PVCpnf(例如SEQ ID NO.93的)，其存在于不对称光杆状菌ATCC43949中。也就是说，PAU_03337的起始密码子开始于PAU_03338的终止密码子末端下游325bp。
这可以通过参考可通过GenBank登录号FM162591.1获得的不对称光杆状菌ATCC43949完整基因组来说明(还参见例如Wilkinson等，BMC Genomics 10，文章号:302(2009)，通过引用并入本文中)，其中效应子基因PAU_03337被注释为如下位于基因组中：补体(3913237..3914247)-即，在核苷酸位置3913237..3914247；并且PAU_03338注释为如下位于基因组中：补体(3914573..3915454)。在这两个基因之间没有发现其他ORF(编码效应子或其他)。
与PVC操纵子相关的另一个PVC效应子基因在本文中称为PVCpnf(例如SEQ IDNO.93)，即PAU_03332(在本文中称为“pnf”)，位于pvc16(PAU_03338)的下游3535bp处。
PVC效应子基因PAU_02095(由于与毒性Rhs毒素基因同源，在本文中称为“Rhs样毒素效应子”)位于存在于不对称光杆状菌ATCC43949中的在本文中称为PVClopT(例如SEQ IDNO.94的)的PVC操纵子的pvc16(PAU_02099)的下游3961bp处。也就是说，PAU_02095的起始密码子开始于PAU_02099的终止密码子末端下游3961bp。
在再一个实例中，基因PAU_02009的PVC效应子(由于预测的作为细胞周期抑制因子/ATP/GTP结合蛋白的功能，在本文中称为“cif”)位于在不对称光杆状菌ATCC43949中发现的相关PVC操纵子(在本文中称为PVCcif)的pvc16(PAU_02008)下游157bp处。
在更进一步的实例中：关于发光光杆状菌TT01的PVC操纵子(在本文中称为PVCunit4操纵子)，PVC效应子基因“pvc17”(例如“plu1651”)位于pvc16(基因“plu1655”)下游104bp处；且关于温度光杆状菌temperata亚种的PVC操纵子Meg1(在本文中称为PVCcif操纵子)，PVC效应子基因“CIF毒素效应子”(例如MEG1 DRAFT_03529)位于相关pvc16基因下游4216bp处。
这些实施例说明编码PVC效应子的基因通常位于PVC操纵子(例如pvc16)的最终基因下游≤5kb的距离内，更通常位于PVC操纵子的最终基因下游≤1kb的距离内。
总之，在这四种菌株中已经鉴定了46种PVC效应子(基于当前可用的序列数据)(参见表1)。这些PVC效应子中的每一种的头50个氨基酸代表(或涵盖)其内源性前导序列，并且发明人已经证明了前导序列可以克隆并融合至多种有效载荷以包装到PVC针状复合物中-参见实施例3和4。因此，PVC效应子(翻译时)包含至少两个主要结构域：前导序列(氨基酸1至50)和实际效应子多肽(氨基酸51至C-末端氨基酸)-其后者在本文中可称为“效应子”(例如“效应子部分”)或“有效载荷”。
尽管光杆状菌基因组序列持续被修改，但是这个PVC效应子基因的综合列表代表了对此类效应子的全面描述，并且是基于目前可获得的最常见(最佳表征的)光杆状菌菌株的序列数据，并且为技术人员提供了对术语“PVC效应子”以及这些PVC效应子的序列(以及如何搜索/挖掘更多的PVC效应子，例如在替代(基因组)序列中)的理解。如上所述，发明人已经发现了PVC效应子蛋白包含前导序列，其对于引导PVC效应子蛋白(例如有效载荷)被包装/负载到PVC针状复合物中是必需的(并且足够的)。
表1
表1中提供的登录号是出于示例性目的提供的，提供了本文所述的PVC效应子的示例性氨基酸序列(或与其具有高相似性)。所述登录号的序列可以通过GenBank(https://www.ncbi.nlm.nih.gov/genbank/)访问。
基因座标签(以“PAU”或“Plu”开始)对应于分配给可通过上述GenBank获得的基因组序列中的效应子的基因座标签。在鉴定所述菌株的基因组内的PVC效应子基因时，本发明的发明人已经指定了以“PAT”(指菌株不对称光杆状菌Thai菌株PB68.1)和“PAK”(指菌株不对称光杆状菌Kingscliff)开始的基因座标签(以与公众可获得的序列的基因座标签一致的方式)。
这个基因座标签在本文中可用于指相应的PVC效应子多肽。
在一个实施方案中，PVC效应子由选自以下的一个或多个基因编码(括号中为编码的PVC效应子蛋白的SEQ ID NO)：PAK_1985(SEQ ID NO:1)、PAK_1987(SEQ ID NO:2)、PAK_1988(SEQ ID NO:3)、PAK_2075(SEQ ID NO:4)、PAK_2077(SEQ ID NO:5)、PAK_2892(SEQ IDNO:6)、PAK_2893(SEQ ID NO:7)、PAK_2894(SEQ ID NO:8)、PAK_3525(SEQ ID NO:9)、PAT_00148(SEQ ID NO:10)、PAT_00149(SEQ ID NO:11)、PAT_00150(SEQ ID NO:12)、PAT_00152(SEQ ID NO:13)、PAT_02308(SEQ ID NO:14)、PAT_02309(SEQ ID NO:15)、PAT_02310(SEQID NO:16)、PAT_02956(SEQ ID NO:17)、PAT_02957(SEQ ID NO:18)、PAT_03171(SEQ IDNO:19)、PAT_03172(SEQ ID NO:20)、PAT_03177(SEQ ID NO:21)、PAU_02009(SEQ ID NO:22)、PAU_02010(SEQ ID NO:23)、PAU_02095(SEQ ID NO:24)、PAU_02096(SEQ ID NO:25)、PAU_02097(SEQ ID NO:26)、PAU_02098(SEQ ID NO:27)、PAU_02230(SEQ ID NO:28)、PAU_02805(SEQ ID NO:29)、PAU_02806(SEQ ID NO:30)、PAU_02807(SEQ ID NO:31)、PAU_03332(SEQ ID NO:32)、PAU_03337(SEQ ID NO:33)、pUL1651(SEQ ID NO:34)、pUL1671(SEQ IDNO:35)、pUL1672(SEQ ID NO:36)、pUL1690(SEQ ID NO:37)、pUL1691(SEQ ID NO:38)、pUL1712(SEQ ID NO:39)、pUL1713(SEQ ID NO:40)、pUL1714(SEQ ID NO:41)、pUL2400(SEQID NO:42)、pUL2401(SEQ ID NO:43)、pUL2514(SEQ ID NO:44)、pUL2515(SEQ ID NO:45)、pUL1649(SEQ ID NO:46)，或其组合。
在一个实施方案中，PVC效应子由选自以下的一个或多个基因编码(括号中为编码的PVC效应子蛋白的SEQ ID NO)：PAU_02009(SEQ ID NO:22)、PAU_02010(SEQ ID NO:23)、PAU_02095(SEQ ID NO:24)、PAU_02096(SEQ ID NO:25)、PAU_02097(SEQ ID NO:26)、PAU_02098(SEQ ID NO:27)、PAU_02230(SEQ ID NO:28)、PAU_02805(SEQ ID NO:29)、PAU_02806(SEQ ID NO:30)、PAU_02807(SEQ ID NO:31)、PAU_03332(SEQ ID NO:32)、PAU_03337(SEQID NO:33)、Plu1651(SEQ ID NO:34)、Plu1671(SEQ ID NO:35)、Plu1672(SEQ ID NO:36)、Plu1690(SEQ ID NO:37)、Plu1691(SEQ ID NO:38)、Plu1712(SEQ ID NO:39)、Plu1713(SEQID NO:40)、Plu1714(SEQ ID NO:41)、Plu2400(SEQ ID NO:42)、Plu2401(SEQ ID NO:43)、Plu2514(SEQ ID NO:44)、Plu2515(SEQ ID NO:45)、Plu1649(SEQ ID NO:46)或其组合。如上所述，这些基因名称对应于可通过GenBank获得的光杆状菌基因组序列中PVC效应子基因的“基因座标签”。PAT和PAK基因座标签由本发明的发明人生成，使得术语与公众可获得的基因组序列的PAU和Plu基因座标签一致。
因此，PVC效应子可以由上述一个或多个基因编码。
在一个实施方案中，PVC效应子由选自以下的一个或多个基因编码(括号中为编码的PVC效应子蛋白的SEQ ID NO)：PAK_02075(SEQ ID NO:4)、PAU_02009(SEQ ID NO:22)、PAU_02096(SEQ ID NO:25)、PAU_02806(SEQ ID NO:30)、PAU_03332(SEQ ID NO:32)、Plu1651(SEQ ID NO:34)、Plu1649(SEQ ID NO:46)或其组合。
在优选实施方案中，PVC效应子由选自以下的一个或多个基因编码(括号中为编码的PVC效应子蛋白的SEQ ID NO)：PAU_02806(SEQ ID NO:30)、PAU_03332(SEQ ID NO:32)、Plu1651(SEQ ID NO:34)、Plu1649(SEQ ID NO:46)或其组合。
PVC效应子可以具有与选自SEQ ID NO:1-SEQ ID NO:46的氨基酸序列具有至少80％序列同一性(优选至少90％序列同一性；更优选100％序列同一性)的序列。例如，PVC效应子可以具有与选自SEQ ID NO:22-SEQ ID NO:46的氨基酸序列具有至少80％序列同一性(优选至少90％序列同一性；更优选100％序列同一性)的序列。
本发明的发明人已经将gogB1(PAU_02806)和Pnf(PAU_03332)PVC效应子的前导序列鉴定为在将(融合的)有效载荷包装到PVC针状复合物中特别有效。在一个实施方案中，PVC效应子由PAU_02806编码(例如具有SEQ ID NO:30的氨基酸序列)。在一个实施方案中，PVC效应子由PAU_03332编码(例如具有SEQ ID NO:32的氨基酸序列)。
在一个实施方案中，PVC效应子包含(或基本上由其组成)选自SEQ ID NO:1-SEQID NO:46(例如SEQ ID NO:22-SEQ ID NO:46)的一个或多个的氨基酸序列，或其组合。例如，PVC效应子可以包含(或基本上由其组成)选自SEQ ID NO:4、SEQ ID NO:22、SEQ ID NO:25、SEQ ID NO:30、SEQ ID NO:32和SEQ ID NO:46的序列。
在一个实施方案中，PVC效应子包含(或基本上由其组成)SEQ ID NO.:4的氨基酸序列。在一个实施方案中，PVC效应子包含(或基本上由其组成)SEQ ID NO.22的氨基酸序列。在一个实施方案中，PVC效应子包含(或基本上由其组成)SEQ ID NO.25的氨基酸序列。在一个实施方案中，PVC效应子包含(或基本上由其组成)SEQ ID NO:30的氨基酸序列。在一个实施方案中，PVC效应子包含(或基本上由其组成)SEQ ID NO:32的氨基酸序列。在一个实施方案中，PVC效应子包含(或基本上由其组成)SEQ ID NO.46的氨基酸序列。
术语“包装”(与术语“反式包装”和“负载”同义使用)意指通过本发明的前导序列(有效载荷与其连接/融合)将有效载荷引导到组装的PVC针状复合物的内部或末端(尖端)中，使得PVC针状复合物随后配置成用于将有效载荷递送(例如注射)到靶细胞中。因此，有效载荷可以包装在PVC针状复合物内，或者可以包装在PVC针状复合物的末端(或尖端)(例如，有效载荷的至少一部分可以在PVC针状复合物的外部)。
术语“有效载荷”(在本文中与术语“弹头”同义使用)意指包装到组装的PVC针状复合物的内部或末端(尖端)中，并且随后递送(例如注射)到(靶)细胞中的分子。在野生型光杆状菌中，有效载荷是PVC效应子(更具体地，所述PVC效应子的效应子部分)，由PVC操纵子的结构基因下游(3')的基因编码(如上所述)。例如，参见图1(D)的模型PVC操纵子，其具有编码腺苷酸环化酶效应子(例如SEQ ID NO.:33)的效应子基因PAU_03337(列为PVCpnf17)；和编码Pnf效应子(例如SEQ ID NO.:32)的PAU_03332(列为PVCpnf 21)。
本发明的前导序列和有效载荷形成“不同于(例如野生型)PVC效应子”(例如由表1中概述的基因之一编码的多肽)的“效应子融合”(或简称“融合”)。例如，效应子融合体可以是嵌合体，其由来自第一PVC效应子的前导序列融合到第二(不同的)PVC效应子(的效应子部分)(优选所述第二PVC效应子的氨基酸51至C末端氨基酸)形成，其中所述第一PVC效应子和所述第二PVC效应子是不同的。效应子融合体可以是嵌合体，其包含(或基本上由其组成)与非PVC效应多肽融合的本文所述的前导序列。效应子融合体可以是嵌合体，其包含(或基本上由其组成)与非光杆状菌属多肽融合的本文所述的前导序列。效应子融合体可以是前导序列-核酸融合体(优选缀合物)，其包含与核酸融合的本文所述的前导序列。
效应子融合体不限于包含与毒性有效载荷融合的前导序列的融合复合体(例如前导序列可以与治疗性有效载荷融合)。因此，如在“效应子融合体”的内容中使用的术语“效应子”意指包装到PVC针状复合物中的有效载荷(其可以提供各种作用，包括产毒和/或治疗效果)。因此，术语“效应子融合体”可与本文中的术语“融合体”互换使用。
术语“效应子融合体”可以与术语“前导序列-有效载荷融合体”和/或“前导序列-有效载荷复合体”同义使用。
可选地或另外地，有效载荷可以不同于PVC效应子蛋白(例如，不同于PVC效应子的氨基酸51至C-末端氨基酸)。例如，有效载荷可以是在野生型光杆状菌属细菌中未发现的多肽或核酸。
对由光杆状菌编码的各种天然PVC效应子有效载荷的大小(例如多肽长度)和结构的分析显示，存在多种不同的PVC效应子长度和结构，证明本发明的PVC针状复合物递送系统的适用性不受目标有效载荷的大小或性质的限制。总之，不需要特定的二级结构、生物物理性质或货物长度，证实PVC针状复合物可以用作通用多功能递送载体。
有效载荷可以是选自多肽(例如多肽有效载荷)、核酸(例如核酸有效载荷)的一种或多种，或其组合。在优选实施方案中，有效载荷是多肽。
多肽有效载荷的实例包括抗体(例如抗MDM抗体)、纳米抗体、肽疫苗(例如酪氨酸酶相关蛋白2(TRP2)肽疫苗)、核因子-κB抑制剂、T3SS有效载荷(例如抑制NF-κB和/或MAPK途径的T3SS有效载荷)、抗凋亡肽(例如BH4)、烟酰胺腺嘌呤二核苷酸醌内部氧化还原酶(Ndi1)、PHOX复合物亚基、肌管蛋白、核酸(优选DNA)修饰酶或其组合。合适的核酸修饰酶的实例包括重组酶(例如Cre重组酶)、转座酶、Cas酶(例如Cas9)和/或Mad7(优选Mad7，更优选Cre重组酶)。有效载荷可以是例如tBid(SEQ ID NO.:109)和/或BaxBH3肽(aa59-73)(SEQID NO.:111)。
具有酶活性的任何多肽可以是有效载荷。
核酸有效载荷可以与本发明的前导序列缀合/交联。例如，无铜点击化学(例如应变促进的叠氮炔环加成(SPAAC))可用于将核酸与前导序列交联。核酸有效载荷的实例包括引物、mRNA、核酸类似物、适体、小干扰RNA(siRNA)、微RNA治疗抑制剂(antimiR)、微RNA治疗模拟物(promiR)、长非编码RNA调节剂、单指导RNA(sgRNA)或其组合。
前导序列可以直接或间接(例如通过间隔区)与有效载荷融合。前导序列可以与有效载荷共价或非共价融合。在优选实施方案中，前导序列与有效载荷共价融合。例如，融合体/效应子融合体可以是(重组)融合蛋白，其包含(或基本上由其组成)与(多肽)有效载荷融合的PVC效应前导序列。
本发明的另一个方面提供了分离的核酸，其包含编码本发明的PVC效应子前导序列的核苷酸序列。本发明的另一个方面提供了分离的核酸，其包含编码本发明的效应子融合体(例如融合体)的核苷酸序列，和任选编码PVC针状复合物的核苷酸序列。
本发明的另一个方面提供了表达载体，其包含：核酸(优选分离的核酸)，所述核酸包含编码本发明的PVC效应子前导序列的核苷酸序列的。本发明的另一个方面提供了表达载体，其包含：核酸(优选分离的核酸)，所述核酸包含编码本发明的效应子融合体(例如融合体)的核苷酸序列，和任选编码PVC针状复合物的核苷酸序列。
本发明的另一个方面提供了包含分离的核酸的宿主细胞，所述分离的核酸包含编码本发明的PVC效应子前导序列的核苷酸序列。本发明的另一个方面提供了包含分离的核酸的宿主细胞，所述分离的核酸包含编码本发明的效应子融合体(例如融合体)的核苷酸序列，以及任选编码PVC针状复合物的核苷酸序列。
术语“核酸”可以与术语“多核苷酸”同义使用。
本发明的另一个方面提供了包含表达载体的宿主细胞，所述表达载体包含编码本发明的PVC效应子前导序列的核苷酸序列。本发明的另一方面提供了包含表达载体的宿主细胞，所述表达载体包含编码本发明的效应子融合体(例如融合体)的核苷酸序列，以及任选编码PVC针状复合物的核苷酸序列。
所述宿主细胞可以是哺乳动物细胞、昆虫细胞、酵母细胞、细菌细胞(例如大肠杆菌)或植物细胞。在优选的实施方案中，宿主细胞是细菌细胞(优选大肠杆菌)。
在一个实施方案中，宿主细胞是光杆状菌属细胞，任选其中光杆状菌属细胞包含与诱导型启动子可操作地连接的PVC操纵子(例如参见实施例3)。PVC操纵子对于光杆状菌属细胞可以是内源性的(例如，PVC操纵子可以是PVCu4)。合适地，光杆状菌属细胞可以从ATCC以登录号ATCC 29999获得。
本发明的序列(例如前导序列和/或核酸序列)包括已经从其天然存在的环境中取出的序列、重组或克隆(例如DNA)的分离物，以及化学合成的类似物或由异源系统生物合成的类似物。
本发明的前导序列和/或多核苷酸可以通过本领域已知的任何方法制备。例如，可以通过在合适的宿主细胞中复制和/或表达来产生大量的前导序列和/或多核苷酸。通常将编码所需片段的天然或合成DNA片段结合到能够引入原核或真核细胞中并在其中复制的重组核酸构建体(通常是DNA构建体)中。通常，DNA构建体将适合于在单细胞宿主(如酵母或细菌)中自主复制，但也可以用于引入并整合到培养的细菌、昆虫、哺乳动物、植物或其他真核细胞系的基因组中。
本发明的前导序列和/或多核苷酸也可以通过化学合成来产生，例如通过亚磷酰胺方法或三酯方法产生多核苷酸，并且可以在商业自动化寡核苷酸合成仪上进行。双链(例如DNA)片段可以通过合成互补链并在适当的条件下将链退火在一起或通过使用DNA聚合酶与适当的引物序列添加互补链而从化学合成的单链产物获得。
应用于前导序列或核酸序列时，在本发明内容中的术语“分离的”表示前导序列和/或多核苷酸序列已从其天然遗传环境中取出，因此不含其他外来或不需要的编码序列(但可以包括天然存在的5'和3'非翻译区，如启动子和终止子)，并且是以适用于基因工程化蛋白质生产系统的形式。此类分离的分子是从其天然环境中分离的分子。
序列同源性
可以使用多种序列比对方法中的任一种来确定同一性百分比，包括但不限于全局方法、局部方法和杂交方法，如，例如，区段逼近方法。确定同一性百分比的方案是本领域技术人员范围内的常规程序。全局方法从分子的开始到结束比对序列，并通过将各个残基对的得分相加并通过施加缺口罚分来确定最佳比对。非限制性方法包括例如CLUSTAL W，参见例如Julie D.Thompson等，CLUSTAL W：Improving the Sensitivity of ProgressiveMultiple Sequence Alignment Through Sequence Weighting,position-specific GapPenalties and Weight Matrix Choice，22(22)Nucleic Acids Research 4673-4680(1994)；和迭代精修，参见例如Significant Improvement in Accuracy of MultipleProtein.Sequence Alignments by Iterative Refinement as Assessed by Referenceto Structural Alignments，264(4)J.MoI.Biol.823-838(1996)。局部方法通过鉴定由所有输入序列共享的一个或多个保守基序来比对序列。非限制性方法包括例如匹配框，参见例如Eric Depiereux和Ernest Feytmans,Match-Box：A Fundamentally New Algorithmfor the Simultaneous Alignment of Seumber Protein Sequences，8(5)CABIOS 501-509(1992)；Gibbs取样，参见例如C.E.Lawrence等，Detecting Subtle Sequence Signals：A Gibbs sampling Strategy for Multiple Alignment,262(5131)Science 208-214(1993)；Align-M，参见例如Ivo Van Walle等，ALIGN-M-A New Algorithm for MultipleAlignment of Highly Divergent Sequences，20(9)Bioinformatics:1428-1435(2004)。
因此，通过常规方法测定序列同一性百分比。参见，例如，Altschul等Bull.Math.Bio.48:603-16，1986以及Henikoff和Henikoff，Proc.Natl.Acad.Sci.USA 89：10915-19，1992。简而言之，使用缺口开放罚分10、缺口延伸罚分1和如下所示的Henikoff和Henikoff(同上)的“blosum 62”评分矩阵(氨基酸由标准单字母代码表示)比对两个氨基酸序列以优化比对评分。
两个或更多个核酸或氨基酸序列之间的“序列同一性百分比”是序列共有的相同位置的数目的函数。因此，％同一性可以计算为相同核苷酸/氨基酸的数目除以核苷酸/氨基酸的总数，乘以100。序列同一性％的计算还可以考虑缺口的数量，以及需要引入以优化两个或更多个序列的比对的每个缺口的长度。序列比较和两个或更多个序列之间的同一性百分比的确定可以使用本领域技术人员熟悉的特定数学算法(例如BLAST)进行。
用于确定序列同一性的比对评分
然后将同一性百分比计算为：
基本上同源的多肽的特征在于具有一个或多个氨基酸取代、缺失或添加。这些变化优选具有次要性质，即保守氨基酸取代(参见下文)和不显著影响多肽的折叠或活性的其他取代；小缺失，通常为1至约30个氨基酸；和小的氨基或羧基末端延伸，如氨基末端甲硫氨酸残基、多达约20-25个残基的小接头肽，或亲和标签。
保守氨基酸取代
碱性：精氨酸、赖氨酸、组氨酸
酸性：谷氨酸、天冬氨酸
极性：谷氨酰胺、天冬酰胺
疏水性：亮氨酸、异亮氨酸、缬氨酸
芳香族：苯丙氨酸、色氨酸、酪氨酸
小的：甘氨酸、丙氨酸、丝氨酸、苏氨酸、甲硫氨酸
除了20种标准氨基酸之外，非标准氨基酸(如4-羟基脯氨酸、6-N-甲基赖氨酸、2-氨基异丁酸、异缬氨酸和α-甲基丝氨酸)可以取代本发明多肽的氨基酸残基。有限数量的非保守氨基酸、不由遗传密码编码的氨基酸和非天然氨基酸可以取代多肽氨基酸残基。本发明的多肽还可以包含非天然存在的氨基酸残基。
非天然存在的氨基酸包括但不限于反式-3-甲基脯氨酸、2,4-亚甲基-脯氨酸、顺式-4-羟基脯氨酸、反式-4-羟基-脯氨酸、N-甲基甘氨酸、别苏氨酸、甲基-苏氨酸、羟基-乙基半胱氨酸、羟基乙基高半胱氨酸、硝基-谷氨酰胺、高谷氨酰胺、哌啶酸、叔亮氨酸、正缬氨酸、2-氮杂苯丙氨酸、3-氮杂苯丙氨酸、4-氮杂苯丙氨酸和4-氟苯丙氨酸。本领域已知用于将非天然存在的氨基酸残基结合到蛋白质中的几种方法。例如，可以采用体外系统，其中使用化学氨基酰化的抑制剂tRNA抑制无义突变。用于合成氨基酸和氨酰化tRNA的方法是本领域已知的。含有无义突变的质粒的转录和翻译在包含大肠杆菌S30提取物和市售酶和其他试剂的无细胞系统中进行。通过色谱法纯化蛋白质。参见，例如，Robertson等，J.Am.Chem.Soc.113:2722，1991；Ellman等，Methods Enzymol.202:301，1991；Chung等，Science 259:806-9，1993；和Chung等，Proc.Natl.Acad.Sci.USA 90:10145-9，1993。在第二种方法中，通过微注射突变的mRNA和化学氨基酰化的抑制剂tRNA在非洲爪蟾卵母细胞中进行翻译(Turcatti等，J.Biol.Chem.271:19991-8，1996)。在第三种方法中，在不存在待替代的天然氨基酸(例如苯丙氨酸)和存在所需的非天然存在的氨基酸(例如2-氮杂苯丙氨酸、3-氮杂苯丙氨酸、4-氮杂苯丙氨酸或4-氟苯丙氨酸)的情况下培养大肠杆菌细胞。将非天然存在的氨基酸结合到多肽中以代替其天然对应物。参见，Koide等，Biochem.33:7470-6，1994。天然存在的氨基酸残基可以通过体外化学修饰转化为非天然存在的种类。化学修饰可以与定点诱变组合以进一步扩大取代范围(Wynn和Richards，Protein Sci.2:395-403，1993)。
有限数量的非保守氨基酸、不由遗传密码编码的氨基酸、非天然存在的氨基酸和非天然氨基酸可以取代本发明多肽的氨基酸残基。
本发明的多肽中的必需氨基酸可以根据本领域已知的程序鉴定，如定点诱变或丙氨酸扫描诱变(Cunningham和Wells，Science 244：1081-5，1989)。生物相互作用的位点也可以通过结构的物理分析来确定，如通过诸如核磁共振、晶体学、电子衍射或光亲和标记的技术结合推定的接触位点氨基酸的突变来确定。参见，例如de Vos等，Science 255：306-12，1992；Smith等，J.Mol.Biol.224:899-904，1992；Wlodaver等，FEBS Lett.309:59-64，1992。还可以从与本发明多肽的相关组分(例如易位或蛋白酶组分)的同源性分析推断必需氨基酸的身份。
可以使用已知的诱变和筛选方法进行和测试多个氨基酸取代，如Reidhaar-Olson和Sauer(Science 241：53-7，1988)或Bowie和Sauer(Proc.Natl.Acad.Sci.USA 86：2152-6，1989)公开的那些。简而言之，这些作者公开了用于同时随机化多肽中的两个或更多个位置，选择功能性多肽，然后对诱变的多肽进行测序以确定每个位置处可允许取代谱的方法。可以使用的其他方法包括噬菌体展示(例如，Lowman等，Biochem.30:10832-7，1991；Ladner等，美国专利号5,223,409；Huse，WIPO公开WO 92/06204)和区域定向诱变(Derbyshire等，Gene 46：145，1986；Ner等，DNA 7:12 7，1988)。
可以使用已知的诱变和筛选方法进行和测试多个氨基酸取代，如Reidhaar-Olson和Sauer(Science 241：53-7，1988)或Bowie和Sauer(Proc.Natl.Acad.Sci.USA 86：2152-6，1989)公开的那些。简而言之，这些作者公开了用于同时随机化多肽中的两个或更多个位置，选择功能性多肽，然后对诱变的多肽进行测序以确定每个位置处可允许取代谱的方法。可以使用的其他方法包括噬菌体展示(例如，Lowman等，Biochem.30:10832-7，1991；Ladner等，美国专利号5,223,409；Huse，WIPO公开WO 92/06204)和区域定向诱变(Derbyshire等，Gene 46：145，1986；Ner等，DNA 7:12 7，1988)。
除非另有定义，否则本文使用的所有技术和科学术语具有与本公开所属领域的普通技术人员通常理解的含义相同的含义。Singleton等，DICTIONARY of MICROBIOLOGY andMOLECULAR BIOLOGY，第20版，John Wiley and Sons，New York(1994)和Hale&Marham，THEHarper COLLINS DICTIONARY of BIOLOGY，Harper Perennial，NY(1991)为技术人员提供了本公开中使用的许多术语的通用词典。
本公开不受本文公开的示例性方法和材料的限制，并且与本文描述的那些类似或等同的任何方法和材料可用于实践或测试本公开的实施方案。数值范围包括限定该范围的数字。除非另有说明，否则分别地，任何核酸序列以5'至3'方向从左到右书写；氨基酸序列以氨基到羧基方向从左至右书写。
本文提供的标题不是对本公开的各个方面或实施方案的限制。
氨基酸在本文中使用氨基酸名称、三字母缩写或单字母缩写来提及。如本文所用，术语“蛋白质”包括蛋白质、多肽和肽。如本文所用，术语“氨基酸序列”与术语“多肽”和/或术语“蛋白质”同义。在一些情况下，术语“氨基酸序列”与术语“肽”同义。在一些情况下，术语“氨基酸序列”与术语“酶”同义。术语“蛋白质”和“多肽”在本文中可互换使用。在本公开和权利要求中，可以使用氨基酸残基的常规单字母和三字母代码。氨基酸的3字母代码符合IUPACIUB生物化学命名联合委员会(JCBN)定义。还应当理解，由于遗传密码的简并性，多肽可以由多于一个的核苷酸序列来编码。
术语的其他定义可以出现在整个说明书中。在更详细地描述示例性实施方案之前，应当理解，本公开不限于所描述的特定实施方案，并且因此可以变化。还应理解，本文使用的术语仅用于描述特定实施方案的目的，并不旨在限制，因为本公开的范围将仅由所附权利要求限定。
在提供数值范围的情况下，应当理解，除非上下文另有明确规定，否则还具体公开了该范围的上限和下限之间的每个中间值，至下限单位的十分之一。在所述范围内的任何所述值或中间值与所述范围内的任何其他所述值或中间值之间的每个较小范围都包含在本公开内。这些较小范围的上限和下限可以独立地包括在该范围内或排除在该范围之外，并且其中任一限值、没有限值或两个限值都包括在较小范围内的每个范围也包括在本公开内，受制于所述范围内的任何具体排除的限值。在所述范围包括限值中的一个或两个的情况下，排除包括限值的那些中的任一个或两个的范围也包括在本公开中。
必须注意，除非上下文另有明确规定，否则如本文和所附权利要求中所用，单数形式“一个(a)”、“一个(an)”和“该”包括复数指代物。因此，例如，提及“一种效应子”包括多种这样的效应子，并且提及“该效应子”包括提及一种或多种效应子及其本领域技术人员已知的等同物，等等。
提供本文所讨论的出版物仅仅是因为它们在本申请的提交日之前的公开内容。本文中的任何内容都不应被解释为承认这些出版物构成所附权利要求的现有技术。
附图简述
现在将参考以下附图和实施例仅通过举例的方式描述本发明的实施方案。
图1显示(A)编码PVC针状复合物的一种PVC操纵子布局(存在于起始基因组的不同区域中的基因簇)的示意图。(B)I、II和III类PVC操纵子布局的示意图。类别中的同源亚基类型显示为具有相似的阴影(以灰度表示)。(C)组装的PVC针状复合物的图示。显示的编号用于将(A)中的基因簇与(C)中的结构中编码的蛋白质的位置相关联(例如，A中的帽'16'簇在(B)的最左侧帽区域中显示为'16')。(D)模型I类PaATCC43949PVCpnf操纵子(例如由SEQ IDNO.:93编码)的图谱，显示了有效载荷区域中的两个效应子基因(Rhs样腺苷酸环化酶和PAU_03332)。
图2显示了基于重叠PCR制备PVC针状复合物表达质粒的克隆程序的概述。PCR片段(具有重叠区域)由不对称光杆状菌ATCC43949(可从ATCC以登录号ATCC 43949获得)的模板gDNA提供，其中相关引物靶向PVC操纵子。
图3显示了PVC针状复合物(例如，由具有上述表达载体的细胞制备)的(体外)样品的透射电子显微照片。PVC针状复合物以独特的“纳米注射器”结构组装，与其作为收缩结构的作用一致。(B)中显示了从高分辨率单粒子cryo-EM断层扫描结构导出的PVC针状复合物的3D渲染模型。
图4显示了(A)在用抗Pnf(immunogold)抗体免疫金染色后包含Pnf有效载荷的PVC针状复合物的透射电子显微照片，证实Pnf-有效载荷毒素与PVC针状复合物(称为PVCpnf)缔合。从编码PVCpnf操纵子的大肠杆菌粘粒克隆的上清液制备PVCpnf针状复合物。使用针对Pnf(TGQKPGNNEWKTGR，SEQ ID NO:96)表位的抗肽抗体来定位有效载荷毒素蛋白。Pnf毒素只能在断裂或收缩的针状复合物的末端检测到，提供了毒素包含在复合物内的证据(箭头)。(B)蛋白质印迹分析证实，如果PVC针状复合物被化学或物理破坏，则仅可以使用抗肽抗体检测Pnf蛋白(毒素)。这些制剂取自PaATcc43949上清液。不能检测澄清的上清液中的Pnf证实所有蛋白质与PVC针状复合物富集制剂缔合。泳道1+5；超声样品，2+6；1M NaCl处理，3+7；1％SDS处理4+8；1M尿素处理。注意PVC针状复合物在1M NaCl中显示出是稳定的。
图5显示了来自5龄烟草天蛾的离体血细胞(昆虫巨噬细胞/嗜中性粒细胞等同物)的冷冻SEM图像，所述烟草天蛾已经注射了由大肠杆菌粘粒克隆异源产生的PaATCC43949PVCpnf针状复合物(纳米注射器)的天然(A)或热灭活(B)富集制剂。注意到对应于PVC针状复合物(纳米注射器)(小箭头)和膜褶皱效应(大箭头)的丰富线性结构，与Pnf有效载荷毒素的作用模式一致，其在对照处理中不存在。比例尺＝50μm。25kV；放大倍数40K(A)和50K(B)。
图6显示了证明与PVC针状复合物接触后的(毒性)细胞表型是由于细胞内毒素递送的实验结果。(A)将负载Pnf的PVC针状复合物注射到昆虫(大蜡螟昆虫幼虫)中，对于给定剂量在15分钟内显示出有效的活性(在实施例中解释)-注意到死亡率/发病率通常与这些死亡/垂死昆虫中的“黑化”免疫应答相关。(B)注射到动物中的对照、变性(通过煮沸)的负载Pnf的PVC针状复合物显示没有活性。(C)纯化的Pnf(有效载荷)，不存在PVC针状复合物(即Pnf未包装到复合物中)，没有显示出对抗动物(左)或HeLa细胞系(右)的活性。(D)递送到HeLa细胞的胞质中的Pnf(有效载荷)-通过含有蛋白质的“BioPorter”脂质体制剂，或通过用适当的质粒转染后的胞内表达(E)-显示出有效的活性/毒性，如通过细胞中的多核化所证明的。(F)-PVCpnf+Pnf对THP1衍生的人巨噬细胞的呼吸速率的影响，如通过刃天青读板测定所测量的。注意到热变性的和空的PVCpnf纳米注射器没有显示出强烈的不利影响。通过注射到Galleria幼虫中来测试这些相同的样品。PVCpnf+Pnf样品在几分钟内显示出超过50％的死亡率(底部两个图中变暗的幼虫)，而注射热变性的和空的PVCpnf的昆虫都保持健康(顶部两个图中没有变暗的幼虫)。
图7显示了(在计算机中)与各种PVC操纵子缔合的一系列内源性有效载荷(毒素)的预测二级结构，证明了多种结构类型。(B)相对于预测的等电点绘制的各种有效载荷(毒素)的氨基酸长度。
图8显示了证实本发明的前导序列(例如具有50个氨基酸)对于将有效载荷蛋白/肽(反式)包装到光杆状菌中表达的PVC针状复合物(纳米注射器)中是必需和足够的。(A)1-6:嵌合效应子蛋白表达构建体(在阿拉伯糖诱导型pBAD30载体中反式表达)的示意图，包括表达Pnf和非天然cre重组酶和Myc标签的那些。C-末端Myc标签表位显示为黑色箭头。(B)使用抗Myc小鼠抗体的蛋白质印迹。样品来自经纯化的PVC(u4)针状复合物(纳米注射器)，其从含有(A)中所示的反式包装表达构建体1-6的染色体工程化的发光光杆状菌TT01过表达。空白pBAD30质粒用作阴性对照，并且没有显示出信号。箭头显示预期产品的正确条带尺寸。
图9显示了前导序列的比对，证明了基于氨基酸性质，前导序列之间存在化学组成共有序列。更具体地，前导序列包含类似的电荷模式，2×带负电荷的区域，每个带负电荷的区域后面是带正电荷的区域[-ve][+ve][-ve][+ve]。
图10显示了(A)来自颗粒制剂(氯化铯梯度和Monolith FPLC制剂，如材料和方法中所述)的PVC针状复合物和有效载荷的蛋白质印迹分析。在[1](pBADPVCpnf，其中纳米注射器的PVC16是FLAG标记的，提供用抗FLAG Ab可检测的PVC16::FLAG)中，可以看到来自“PVCPnf”(具有Pnf有效载荷的PVC针状复合物)的标记帽蛋白的信号，证实了纯化级分中存在PVC针状复合物。在[2](pBADPVCpnf+Cre::Myc，用抗Myc Ab可检测，Cre具有Pnf前导序列的N-末端融合，例如SEQ ID NO.:78)中，来自Myc标记的有效载荷蛋白的信号在与(1)相同的样品中大量包装，证实了纯化的PVC针状复合物(纳米注射器)中存在Cre有效载荷。在[3](PVCU4+Cre::Myc，用抗Myc Ab可检测，Cre具有Pnf前导序列的N-末端融合，例如SEQ IDNO.:78)中，探测不同的PVC针复合底盘(“PVCU4”)纯化的Myc标记的Cre，揭示了包装的(包装的Myc标记的Cre)相应条带。为清楚起见，这在印迹中突出显示。(B)PVC针状复合物的透射电子显微照片，显示了野生型(具有Pnf有效载荷)PVC针状复合物和具有非典型(非天然)重组酶(Cre)有效载荷的PVC针状复合物在任何测试的底盘中都不影响PVC针状复合物的形态，确保它们不会异常组装。图10(C)提供了(A)的数据的附加/补充数据。更详细地，(C)通过Cre重组酶(反式)包装到大肠杆菌中表达的纯化PVCpnf中的蛋白质印迹分析提供了更多证据。蛋白质印迹表明，对于给定量的抗FLAG抗体蛋白质信号(由于PVC16::FLAG的结合，纳米注射器的特异性探针)，检测到高得多含量的Cre有效载荷(使用抗myc标签抗体)。数字表示2倍稀释。注意到稀释后，来自纳米注射器的抗FLAG信号丢失，而有效载荷在大多数泳道中保持强烈。CsCl表示通过氯化铯密度梯度离心纯化。“Mon”表示样品另外通过“Monolithic”柱进行阴离子交换。“洗脱后”、“相间”、“亚相间”表示其中从纯化过程检测到信号的液体级分。D-反式包装到大肠杆菌中的PVCpnf中的Cre的蛋白质印迹分析。在纳米注射器-有效载荷复合物纯化后，探测有效载荷的结合的“Myc”标签(C-末端融合)。颗粒制备物的蛋白质印迹分析证实所有四种前导序列都可以有效地反式包装外源性Cre酶。E-A系统发生树，证明所示例的前导序列整体分布良好，因此处于或接近最大程度地顺序多样化(参见实施例4.2)。
图11显示了用抗FLAG和抗myc抗体同时探测的在没有(1)和有(2)来自分开的质粒的(myc标记的)Pnf的伴随表达的情况下表达的PVC针状复合物的蛋白质印迹分析。在标记为1的泳道中，在大肠杆菌内不存在“有效载荷质粒”(编码与前导序列连接的有效载荷蛋白的表达质粒)的情况下表达和纯化PVC针状复合物(纳米注射器)。这导致仅对应于注射器(PVC针状复合物)本身上存在的FLAG标签的条带。对于泳道2，采用相同的方法，但使用还包括携带标记的有效载荷(Myc-Pnf)的(分开的)质粒的培养物。可以看到对应于FLAG和Myc标签的条带，证实Pnf有效载荷的存在(1和2内的四个泳道是来自氯化铯梯度的简单不同的纯化级分)。
图12显示了发光光杆状菌TT01 PVCu4过表达菌株中反式包装实验的蛋白质印迹分析。结果证明了myc标记的Pvc17(pLU1651whole::Myc)的反式包装。
图13显示了发光光杆状菌TT01 PVCunit4过表达菌株中的反式包装实验的进一步蛋白质印迹分析(如实施例中所解释的)。结果证明了使用Pnf的前导序列(PAU_03332前导序列)的Myc标记的Pvc17(Plu 1651::Myc)和单独的Myc标签的反式包装，并且前导序列是必需的。(A)泳道1显示了与Myc标签融合的前导序列(PAU_03332::Myc)的包装；泳道3显示了不存在前导序列时缺乏包装(仅Myc未被包装)；泳道4显示不存在前导序列时缺乏HvnA(天然效应子)的包装；泳道6显示了Myc标记的PAU_03332::Plu1649的包装，即来自PAU_03332的前导序列(即PAU_03332的氨基酸1-50)和来自Plu1649的效应子(即氨基酸51-C-末端)的嵌合体。泳道1和6中的高强度条带证明了Pnf(PAU_03332)前导序列在包装有效载荷方面特别有效)。(B)泳道1显示了使用抗Myc抗体蛋白质印迹的具有C-末端Myc标签的Plu1651的包装。
图14显示了进一步的蛋白质印迹分析，证明了使用PAU_02806(GogB)前导序列(第二泳道，不包括阶梯泳道)非常高水平的的Myc标记的Pnf(PAU_03332::Myc)的反式包装。第一泳道证明了Plu1649前导序列用于包装PAU_03332效应子(Myc标记的Plu1649::PAU_03332)的用途。由于第二泳道中条带的相对强度，条带看起来较弱。该实验涉及50mL培养物的过滤灭菌，加入8M终浓度的尿素以分解PVC。从10mL上清液中收集样品。
图15显示了进一步的蛋白质印迹分析，证明了如图13中所述的具有C-末端Myc标签的Plu1651(pvc17)反式包装到由光杆状菌属表达的PVCunit4中。Raw代表来自上清液的颗粒制备物，Be、Be2和IP代表来自氯化铯梯度纯化的不同“切割”。
图16(A)提供了Cre在(实施例6的)小鼠类器官实验中的作用机制的图解说明，以及阳性对照(TAM)如何促进Cre活化。白色箭头显示表达tdTom荧光报告基因的细胞的位置。B-证明通过从大肠杆菌表达和纯化的PVCpnf将活性反式包装的Cre重组酶递送到鼠胆管类器官中。白色圆圈显示表达荧光报告基因的细胞组的位置。上图显示了通过光学显微镜获得的图像的直接灰度转换。下图显示了具有阳性细胞的假颜色增强的相应图像，提供其以简单地帮助识别在前一灰度转换内受影响的细胞和周围未受影响的细胞之间的差异。
图17显示了具有和不具有有效载荷(Cas9样蛋白MAD7)的纳米注射器表达的斑点印迹分析。在诱导前(T1)观察到IPTG诱导型MAD7的一些泄漏表达，这是该表达系统常见的。如预期的，在任何时间点都没有来自仅PVC样品的Myc信号，并且MAD7信号在～24小时内的整个表达过程中生长。如别处所述，通过超速离心纯化后维持强Myc信号，表明蛋白质结合到纳米注射器底盘系统中。FLAG信号在MAD7样品中是稳健的，并且如预期在诱导后发生并且在纯化后持续，因为这个启动子系统具有减少的泄漏表达。得出结论，纳米注射器和MAD7在表达方面彼此相容，并且迄今为止测试的最大蛋白质MAD7可以包装到纳米注射器系统中。
图18显示了蛋白质斑点印迹分析，证实了促凋亡tBid蛋白结构域和BaxBH3肽(两者均具有与N-末端融合的SEQ ID NO.:78的前导序列)反式包装到纯化的从大肠杆菌表达的PVCpnf(7&8)中。显示了通过2种不同方法(5&6)纯化的具有其同源毒素“Pnf”的纳米注射器作为阳性对照。图底部的印迹代表与上述图中的7&8中的相同实施例。这些印迹由相同构建体的另一次纯化制备，证明了纯化的再现性。该实验证明了可以成功地制备“tBid蛋白结构域和BaxBH3肽”包装的样品(纳米注射器)，例如用于实施例9中的细胞凋亡递送测定中。
图19(A)显示了来自仅暴露于包装的纳米注射器20分钟的细胞的TUNEL染色显微镜分析。第一(左)条＝DNA酶I处理的细胞(+对照)；第二条＝无DNA酶I或纳米注射器处理(-对照)；第三条＝将细胞暴露于包装有tBid(通过与N-末端融合的SEQ ID NO.:78的前导序列)的纳米注射器；第四(右)条＝将细胞暴露于包装有Bax_BH3结构域(通过与N-末端融合的SEQ ID NO.:78的前导序列)的纳米注射器。B-如实施例9中所述的代表性显微照片，其显示了用纳米注射器和对照处理后PBMC的TUNEL染色。在进行TUNEL染色以确定凋亡应答之前，用tBID、Bax加载的纳米注射器以及阳性(DNA酶I处理的细胞)和阴性(无DNA酶I处理)对照在室温下处理PBMC 20分钟。在原始(非灰度)显微照片中：凋亡反应阴性的细胞显示蓝色或浅棕色染色。蓝色染色(甲基绿)或浅棕色染色指示不存在凋亡信号的健康细胞。深棕色染色表明细胞经历凋亡。
实施例
材料和方法
克隆
使用本领域已知的标准分子技术制备编码PVC针状复合物的质粒。简而言之，来自不对称光杆状菌ATCC43949的基因组DNA(可依据登录号ATCC43949从ATCC获得)用于PCR(使用适当的引物)以扩增PVC操纵子的多个(例如四个)重叠区域。采用重叠/延伸PCR来制备完整操纵子，并融合(再次使用重叠PCR)到如图1中详述的适当表达载体中(使用SEQ ID NO:101-SEQID NO:106的引物)。
简而言之：制备覆盖PVC操纵子(例如SEQ ID NO:93)的四个重叠PVC片段(分别用SEQ ID NO:101(F1)和SEQ ID NO:105(R1)、SEQ ID NO:102(F2)和SEQ ID NO:106(R2)、SEQID NO:103(F3)和SEQ ID NO:107(R3)以及SEQ ID NO:104(F4)和SEQ ID NO:108(R4)的引物产生)。在所需插入位点切割靶克隆载体。然后通过重叠PCR(使用SEQ ID NO:101和SEQID NO:108的引物)组装这5个DNA片段，并将所得片段连接到克隆载体中。将产物转化到实验室大肠杆菌中并用载体标志物选择(例如由于氨苄青霉素抗性)回收。
如本领域已知的，操纵子通常可操作地连接诱导型启动子(例如阿拉伯糖诱导型和/或IPTG诱导型)。这通常通过克隆到pBAD家族质粒(可通过阿拉伯糖诱导)(Invitrogen，目录号:V43001)和pVTRa(可通过IPTG诱导)(Biomedal，S.L.)载体中来实现(尽管相容表达载体系统的任何组合都应该是足够的)。
PVC针状复合物可以独立于有效载荷(毒素)表达，反之亦然。分开的表达载体(例如具有不同的诱导型启动子)可以分别携带PVC针状复合物和有效载荷。
在大肠杆菌中表达(例如实验室规模表达)/纯化PVC针状复合物
从大肠杆菌表达菌株(用适当的表达载体/粘粒转化)的1L培养物中纯化PVC针状复合物的典型方法如下：
1-通过从平板中挑取菌落并接种100mL LB培养基来制备细菌(用PVC Needle复合物表达载体转化)的过夜培养物。
培养物在37℃下振荡生长。
a.通常，培养基可以常规补充0.2％d-葡萄糖，以帮助抑制遗传构建体以获得最佳细胞健康。
b.培养基还补充有用于维持表达(PVC Needle复合物)载体的相关抗生素。如果还使用有效载荷载体，则还提供该载体的相关抗生素。
2-第二天，通过以1:100的比例稀释接种1L烧瓶过夜培养物。1L烧瓶的培养基与过夜培养基相同，但通常不含葡萄糖。
3-培养物生长至大约指数中到后期(～0.8的OD600nm)，此时诱导质粒。
a.对于PVC针状复合物(纳米注射器)质粒，通常加入0.2％阿拉伯糖以诱导表达。对于有效载荷质粒(编码有效载荷的质粒，如Pnf)，IPTG浓度通常可以基于每个蛋白质进行优化，并且0.1mM的典型起始数字是优选的。
4-在诱导后将培养物返回培养箱并在18℃下培养直至第二天。
5-通过在适当的离心机/瓶/转子中5000×g持续30分钟离心收获培养物。
6-然后裂解细胞沉淀物以释放PVC针状复合物(纳米注射器)。
a.可以使用以下裂解方法：
(i)溶菌酶温育过夜。(ii)用针式超声波仪超声处理(首先用溶菌酶处理或不用溶菌酶处理)。(iii)破碎器/均化器。
7-任选地，可以将DNA酶和蛋白酶抑制剂添加到裂解物中。
8-通过在高速离心机中以50,000×g，4℃，离心20分钟除去细胞碎片。
9-通过100,000kDa MWCO离心柱浓缩裂解物以减少体积并除去小蛋白质。一旦体积降至可控制的体积，离心数次，用合适的样品缓冲液，如TM(20mM Tris-HCl，8mM MgCl2，20mM Tris-HCl；2pH 7.4)替代截留液以进行渗析。
通过氯化铯密度梯度纯化的后续过程如下：
1.如下制备CsCl密度溶液：
(a)H2O中的1.7g/mL CsCl；(B)H2O中的1.5g/mL CsCl；(C)H2O中的1.45g/mL CsCl
2.然后在超速离心管中设置梯度(从管的底部到顶部)，如下：
(1)(管底部)-2mL密度，1.7CsCl；(2)-3ml密度，1.5CsCl；(3)-3mL密度，1.45CsCl；(4)(管顶部)-TM缓冲液中的样品。适当地，将每个密度小心地施加到管的侧面，使得边界不与先前的密度层混合。
3.然后将平衡的管在SW40Ti摆动桶式转子中在4℃下以35,000rpm(相当于155,000×g)超速离心2小时。
4.正确的梯度级分将是刚好在出现的“蓝-白”晕上方的区域。通过用注射器和针穿刺管来提取级分。
5.可以以这种方式获得良好纯度的PVC针状复合物，并在4℃下储存在缓冲液中。适当地，渗析回TM缓冲液中以远离CsCl。
在CsCl梯度纯化之后或代替CsCl梯度纯化，可以通过Monolith阴离子交换色谱法提取PVC，如下(注意所有步骤可以用蠕动泵或注射器装置手动进行，或通过F/HPLC进行)：
1.除非已经进行，否则将样品提取物渗析到具有低浓度盐(20mM NaCl)的结合流动相(通常为TM缓冲液)中。
2.根据制造商的指导对柱进行平衡，简要地：
a.至少5个柱体积(CV)的dH2O；
b.至少5CV的结合缓冲液(TM，低盐)；
c.至少5CV的洗脱缓冲液(TM，高盐，>＝1M NaCl)；
d.再次至少10CV的结合缓冲液。
3.以低流速(1-2mL/min)将样品施加到柱上
4.用至多200mM含NaCl的TM缓冲液洗涤柱。
5.用含有1M NaCl的TM缓冲液洗脱(或者，如果使用FPLC机器，则使用梯度洗脱)。
6.PVC针状复合物存在于洗脱级分中。如果使用级分收集器，则可能需要随后的SDS-PAGE或类似操作来鉴定正确的级分。
(例如步骤2的)柱是CIMmultus(TM)季胺阴离子交换柱(BIA Separationsd.o.o.)。例如，CIMmultusTMQA-1，其是具有1.3μm通道尺寸和1mL柱体积的整体柱。
或者，可以使用DEAE(弱阴离子交换剂)柱。
或者，为了与光杆状菌表达系统一起使用，可以以及/代替细胞沉淀物从上清液中纯化PVC针状复合物，使用如下添加/修改：
1.从上述标准方案收获细胞后，将上清液转移到pyrex瓶中，并且如果需要，可以任选地通过100,000MWCO柱浓缩。
a.可以任选地添加DNA酶(0.25U/mL)和蛋白酶抑制剂。
2.加入NaCl至终浓度为0.5M，并且还加入80g/L的PEG6000。将溶液在4℃下混合过夜。
3.将溶液在8000×g，4℃下离心30min以沉淀PEG6000。
4.将沉淀物重悬于小体积(～5mL)TM缓冲液(或类似物)中，并在室温下振荡孵育2小时。
5.通过以13,000×g离心10min来沉淀，并将上清液收集到新管中。进行选择的纯化方法。
用于纯化PVC针状复合物的其他方法已经在其他地方描述，例如在Yang等(JBacteriol.2006 Mar；188(6)：2254-2261)中，其通过引用并入本文。
用于发光光杆状菌TT01 PVCunit4(由基因plu1667-plu1652编码的底座)的阿拉
伯糖诱导型过表达菌株的构建
使用染色体重组将选择的PVC(操纵子)(作为实例，本文使用了编码PVCunit4针状复合物的操纵子)置于阿拉伯糖诱导型转录启动子的控制下来制备过表达PVC针状复合物的光杆状菌属菌株。然后用效应子表达质粒(例如，基于阿拉伯糖诱导型表达载体pBAD30)对重组菌株进行遗传转化，以简单地通过添加阿拉伯糖来促进PVC针状复合物过表达、PVC效应子表达、PVC效应子反式包装和整个复合物的分泌。
重组光杆状菌PVC过表达菌株的构建
使用来自发光光杆状菌DJC(也称为菌株TT01)的基因组DNA作为模板，使用引物PVCpromF(5'-TATCATATGTCTACAACTCCAGAACAAATTGC TG-3'，SEQ ID NO:97)和PVCpromR(5'-ATCTCTAGAACAGATATTCCAGCCAGC-3',SEQ ID NO:98)，来扩增PVCunit4的启动子区。合适的发光光杆状菌菌株可依据登录号ATCC 29999从ATCC获得。用NdeI和XbaI消化PCR产物，并使用大肠杆菌DH5αλ-pir(Biomedal S.L.)作为载体菌株，通过连接引入自杀载体pCEP中(ThermoFisher，目录号:V04450)。将所得质粒转移到大肠杆菌供体菌株S17.1λ-pir中(Biomedal S.L.)，用于缀合到光杆状菌属中。简而言之，将供体菌株和发光光杆状菌DJC的利福平抗性(RifR)分离株的过夜培养物在补充有10mM MgSO4的LB中稀释，并生长至指数中期(OD600～0.5)。然后，收获3ml的每种培养物，洗涤两次并重悬于100μl补充有10mM MgSO4的LB中。将80μl发光光杆状菌DJC RifR与20μl供体细菌混合(导致受体与供体比率为4:1)并置于补充有0.1％丙酮酸和10mM MgSO4的LB琼脂平板的中心。将板在30℃下孵育过夜，并在1.5ml LB中收获所得生长物。将等分试样涂布于含有利福平(50μg/ml)和氯霉素(25μg/ml)的平板上以选择反式缀合物，并将板在30℃下孵育3天。使用引物ParaINF(5'-GGCGTCACACTTTGCTATG-3'，SEQ ID NO:99)和tPVCpR(5'-TCGGTGGCAGTAAATTGTCC-3'，SEQID NO:100)将可能的反式缀合物重新划线并通过PCR确认。
来自光杆状菌的PVC针状复合体的过表达和纯化
将发光光杆状菌DJC PVCunit4::pCEP的过夜培养物在补充有氯霉素(25μg/ml)的2×250mLLB中稀释，并在28℃、180rpm下孵育。2-3h后，加入阿拉伯糖(0.2％)，并将培养物返回培养箱中另外26h。通过离心(7000g，30min)沉淀细胞并收集上清液。将DNA酶I以0.25U/ml的浓度加入上清液中以降解任何胞外DNA。在室温下孵育30min后，加入聚乙二醇8000(8％)和NaCl(0.5M)以沉淀蛋白质。将上清液在4℃下搅拌孵育过夜。然后通过在4℃下以8000g离心30min来收集沉淀的蛋白质。将沉淀物重悬于8ml TM缓冲液(20mM TrisHCl，20mM MgCl2，pH7.4)中，并在室温下轻轻振荡孵育2h。通过以13000g离心10min来除去任何剩余的碎片，并将含有PVC针状复合物的上清液施用于CsCl密度梯度，并在Beckmancoulter Optima L-90K或XPN-80K超速离心机中以35000rpm离心2h。CsCl密度梯度通过从管的底部分别以p＝1.7(2mL)、1.5(3mL)和1.45(3mL)分层含有CsCl的TM缓冲液来制备。收集含有PVC针状复合物的级分，并使用UltraceMOOK装置(Amicon)除去CsCl，并将缓冲液更换为TMS(20mM TrisHCl，8mM MgSO4，pH7.4)。使用CIMmultusTM季胺2μm孔阴离子交换柱(BiasParations)进一步纯化PVC针状复合物。用含有200mM NaCl的TMS缓冲液洗涤柱，并在含有1M NaCl的TMS中洗脱PVC针状复合物。使用UltraceMOOK装置通过缓冲液交换除去NaCl，并将样品施加到CIMmultusTMDEAE 2μm孔柱(BIAseparations)上进行最终纯化。将柱在含有200mM NaCl的TMS中洗涤，并将样品在含有500mM NaCl的TMS中洗脱。
可以在有和没有(例如，因为PVC针状复合物似乎从活细胞分泌，并且可以收集在上清液中)细胞裂解(以释放PVC针状复合物)的情况下进行这一点。
透射电子显微镜
对于透射电子显微镜(TEM)，使用涂覆有细碳层的聚乙烯醇缩醛覆盖的300目铜网格作为蛋白质级分的衬底。优选的水性负染剂是3％甲胺钨酸盐。将涂覆的网格在使用前立即暴露于UV光16h，以确保衬底的适当润湿。将10μl液滴施加到TEM网格上，并使蛋白质沉降5min。用滤纸从网格边缘吸收液体，并立即用10μl滤过的负染剂替换。用滤纸部分地除去液滴，并且使网格充分风干，然后用在80kV下运行的JEOL1200EX透射电子显微镜(JEOL，东京，日本)观察它们。
BioPORTER测定和肌动蛋白应力纤维分析。
对于BioPORTER测定(Genlantis)，将80μl纯化的野生型和突变体Pnf蛋白(500μgml-1)或PBS作为阴性对照加入一个BioPORTER管(Genlantis)中并重悬于920μl DMEM中。将样品加入在6孔板中生长的HeLa细胞中并孵育4h。用新鲜的完全培养基替换BioPORTER/蛋白质或PBS混合物，并将细胞孵育20-48h。为了使细胞形态和肌动蛋白细胞骨架可视化，将细胞在4％PBS-甲醛中固定15min，用0.1％Triton X-100透化，并用四甲基罗丹明B异硫氰酸酯(TRITC)-鬼笔环肽(Sigma)和DAPI二盐酸盐(Sigma)染色。用LSM510共聚焦显微镜(Leica)获取图像。
实施例1
PVC针状复合物的克隆和表达
发明人已经成功地从宿主细菌发光杆状菌属切除(克隆)所需的表达基因(例如，其包含在SEQ ID NO:93、SEQ ID NO.:94和/或SEQ ID NO:95内)，并且已经设计了如上所述的在实验室大肠杆菌中可靠的、可扩展的表达系统。已经证明了在分开的质粒上的反式表达使得能够将有效载荷(例如Pnf)结合到注射器中，从而形成多质粒(模块化)平台。
从大肠杆菌纯化后，电子显微镜分析证明了纯化的PVC针状复合物保留正确的“纳米注射器”结构(参见图3)。此外，PVC针状复合物在纯化后保持与有效载荷(例如Pnf)正确缔合(参见图4)，证明发明人已经成功制备了具有用于将有效载荷递送至细胞的正确结构的PVC针状复合物(纳米注射器)。
此外，电子显微镜分析证明了纯化的复合物适当地定位于细胞的细胞表面，并且具有Pnf有效载荷的PVC针状复合物诱导效应子(PVC)与假定机制一致的表型(褶皱)-参见图5。
实施例2
2.1证明PVC针状复合物通过效应子的胞内递送发挥作用
如下将多肽Pnf鉴定为PVC效应子。这在不对称光杆状菌ATCC43949完整基因组-GenBank登录号：FM 162591.1内得到了鉴定。
鉴定了PVC操纵子(不对称光杆状菌ATCC43949 PVCpnf操纵子，其具有SEQ ID NO:93的序列)的最终基因，即pvc16(例如PAU_03338)。PVC基因座的pvc16基因的位置示于图1(A)、(B)和(D)中。鉴定了pvc16的3'不远处(例如在pvc16下游约5kb内)的ORF-一个这样的ORF(PAU_03332)在pvc16下游3535bp。由该推定的效应子ORF编码的多肽(具有SEQ ID NO.:32的序列)的预测功能通过BlastP和HHPRED(https://toolkit.tuebingen.mpg.de/#/tools/HHPRED)的组合获得。然后可以基于与已知细菌毒素(例如来自大肠杆菌的CNF1家族)的直接同源性将该ORF指定为PVC效应子。
然后根据实施例1制备负载Pnf的PVC针状复合物。
发明人已经证明了这些包装的(例如负载的)PVC针状复合物发挥与其携带的货物的来源一致的细胞效应。举例来说，暴露于负载有细胞骨架毒素Pnf的PVC针状复合物的细胞和整个昆虫动物以与细胞骨架毒性一致的方式经历细胞死亡。
通过注射10μl上清液来进行注射实验(注射到昆虫幼虫中)，所述上清液在大肠杆菌培养物的过夜培养物(通常为1L)离心(沉淀)后提供，所述大肠杆菌培养物携带编码具有Pnf的PVC针状复合物(PVCPnf)的粘粒克隆-例如由SEQ ID NO.:93编码的PVC，其包装有SEQID NO.:32的PVC效应子。
证明PVC针状复合物是由于Pnf有效载荷的胞内递送(例如注射)而导致表型的原因，只有当相同的蛋白质(Pnf)提供另一种途径以进入细胞胞质(表达质粒的转染和表达，或通过含有蛋白质的脂质体制剂的电导)时，才能重建毒性作用-参见图6。相反，变性(通过煮沸)PVC针状复合制剂、覆盖在组织培养细胞上的毒素蛋白或注射到整个动物中的毒素蛋白显示没有活性。
2.2.毒性效应子酶Pnf递送到培养的人巨噬细胞中的证据
为了补充上述数据，发明人进行了另外的实验，提供了将毒性效应子酶Pnf递送到培养的人巨噬细胞中的更多证据。
概念：发明人在培养的人THP1衍生的巨噬细胞上测试了从(反式)包装有天然Pnf毒素的大肠杆菌表达和纯化的PVCpnf。与Pnf毒素在昆虫模型中的致死作用不同，先前的脂质体介导的Pnf蛋白转染实验表明了在人Hela细胞中更精细的表型。在那些实验中，细胞在24h时显示出肌动蛋白应力纤维形成，并且在48h时显示出多核化。因此，发明人使用刃天青比色测定测试了容纳/包装有pnf PVC效应子的纯化PVCpnf(纳米注射器)对巨噬细胞呼吸速率的影响。
方法：
刃天青测定背后的背景。研究蓝色化合物刃天青用于测定PVC对巨噬细胞(M0)的活性的测定。刃天青在细胞线粒体中代谢还原，产生粉红色和高荧光化合物试卤灵。PVC对巨噬细胞代谢的影响可以通过将刃天青引入培养基中来确定。受PVC影响的巨噬细胞的数量可以通过将测量的荧光与细胞密度优化曲线的荧光进行比较来推断(参见Czekanska，Methods in Molecular Biology，2011，740，27-32，通过引用并入本文中)。
刃天青用于THP1衍生的巨噬细胞的优化。在不同的接种密度下评估巨噬细胞在18h内的代谢，以确定使用PVC进行该测定的最佳细胞密度。将30mL THP-1细胞培养物以1000rpm沉淀4min，然后重悬于2mL RPMI培养基(还含有10％FBS(v/v)和2mM L-谷氨酰胺)中。使用细胞血细胞计数器计数细胞，然后在培养基中稀释至2×106细胞mL-1的密度。然后在涂布前立即用佛波醇12-肉豆蔻酸酯-13-乙酸酯(PMA)活化THP-1细胞。将200μL细胞一式四份涂布在96孔板中，并进行2倍连续稀释直至达到1.5625×103细胞mL-1的最终细胞密度。将125μL起始细胞稀释液一式四份涂布在同一板上，进行5倍连续稀释，直至达到0.32×103细胞mL-1的细胞密度。还制备了四个空白孔，其含有RPMI和PMA。将板在37℃、5％CO2下孵育48h。从孔中吸出培养基并用新鲜RPMI替换，并将巨噬细胞再孵育24h。将刃天青片剂(VWR)溶解于RPMI(12.5mg/mL)中，并快速连续地向每个孔中加入10μL(孔浓度为1.25mg/mL)。在板读数器上每30分钟测量产生的荧光，持续18h(激发：530-570nm，发射：580-620nm，保持在37℃和5％CO2下)。然后确定随时间的最佳细胞密度以与PVC一起使用。
使用测定进行PVC测试。将稀释至1.25×105mL-1THP-1细胞活化并接种于96孔板中，其中孔含有100μL细胞，最终孔密度为1.25×104细胞mL-1。还一式四份制备了空白孔，其含有细胞而不含PVC样品，以及仅含有培养基和PMA的孔。将板在37℃、5％CO2下孵育48h。然后用新鲜RPMI替换培养基，然后加入10μL每种PVC样品。将板再孵育24h，然后向每个孔中加入10μL刃天青(12.5mg/mL)，并且每30min测量荧光，持续18h(激发：530-570nm，发射：580-620nm，保持在37℃和5％CO2下)。
结果：图6F显示了用PVCpnf+Pnf挑战确实降低了巨噬细胞的呼吸速率，而热变性的或空的PVCpnf纳米注射器没有强烈的副作用。然而，没有添加样品的对照细胞仍然显示出最佳的呼吸速率。对巨噬细胞的影响与昆虫注射毒性测定相关。在这种情况下，两种PVCpnf+Pnf制剂对超过一半的昆虫群显示出致死性，而注射热变性的和空的PVCpnf的昆虫都保持健康。
实施例3
证明前导序列负责有效载荷包装到PVC针状复合物中
令人惊讶地，发明人已经发现了，优选在有效载荷(毒素)蛋白的N-末端上，提供“前导”肽序列可以将有效载荷引导至PVC复合物并允许(例如触发)将有效载荷包装到PVC针状复合物中。发明人已经证明了PVC效应子蛋白的氨基酸残基1-50是/包含前导序列。
为了证明这一点，制备了表达构建体(在染色体工程化的发光光杆状菌TT01中过表达)，其中前导序列(N-末端氨基酸残基1-50)被消融，使得由Plu1649表达的有效载荷(在图中称为“hvnA”，并且具有SEQ ID NO.:46的序列)(出于检测目的为Myc标记的)不存在前导序列(参见图8A-构建体1)。在表达(有效载荷和PVC针状复合物两者)和分离PVC针状复合物(并在凝胶上运行其组分，其包括任何包装的有效载荷)后，通过蛋白质印迹分析在PVC针状复合物内未检测到(Myc标记的)Plu1649(“hvnA”)，证明有效载荷(不存在前导序列)未包装到复合物中(参见图8B，泳道1)，并且因此不与分离的复合物缔合。然而，对于保留前导序列的hvnA，观察到成功的包装，参见泳道2(注意到由于泳道3的条带的相对强度，条带看起来弱)。
令人惊讶地，具有来自不同(非hvnA)PVC效应子的前导序列(即对应于来自PAU_03332效应子的N-末端氨基酸残基1-50)的hvnA(参见图8A，构建体3)被正确包装到复合物中，并且在分离/纯化后保持与PVC针状复合物缔合，如通过myc标记的hvnA的蛋白质印迹检测所证明的(参见图8B，泳道3)。因此，发明人已经证明了‘PAU_03332’前导序列(其与不同的有效载荷Pnf缔合)的令人惊讶的用于包装hvnA有效载荷(即与PAU_03332的有效载荷不同的有效载荷)的能力。这证明了交换PVC效应子的前导序列的能力，允许使用最佳前导序列(具有最佳包装活性)进行包装。
实施例4
4.1证明前导序列指引非典型/外源有效载荷的包装(到PVC针状复合物中)
在本发明的意想不到的技术效果中，发明人已经发现了将本文所述的前导序列与外源(非光杆状菌属)多肽(优选地在N-末端)融合允许将所述外源多肽包装到PVC针状复合物中，其中外源多肽在分离/纯化时保持与PVC针状复合物缔合。举例来说，参见图8B(泳道4)，其证明了与前导序列融合时，非光杆状菌‘Myc’多肽(<10kDa)包装到PVC针状复合物中，以及泳道6，证明了大得多的非光杆状菌“Cre-重组酶”多肽(32kDa)同样可以适当地包装到PVC针状复合物中。
发明人对由光杆状菌编码的各种天然PVC效应子有效载荷的大小(例如多肽长度)和结构进行了深入分析(参见图7)，其显示了多种不同的长度和结构，证明了本发明的PVC针状复合物(纳米注射器)递送系统的适用性不受目的有效载荷蛋白的大小或性质的限制。总之，不需要货物特定的二级结构、生物物理性质或长度，证实PVC针状复合物(纳米注射器)底盘可用作通用多功能递送载体。
此外，外源多肽的这种包装不依赖于所选择的PVC针复合底盘，例如已经使用“PVCpnf”底盘(SEQ ID NO.:93)和“PVC U4”(例如PVCunit4)底盘(对于光杆状菌过表达菌株是内源的)两者完成(参见图10A)。重要的是，发明人已经证明了在任一底盘中包装外源有效载荷不影响PVC针状复合物的形态，确保它们不会异常组装(参见图10B)。
在本文所示的数据中，有效载荷蛋白在分开的遗传构建体上以“反式”提供。前导序列令人惊讶地足以靶向这些分开合成的蛋白质以包装到PVC针状复合物载体中(参见图11)。底盘(PVC)基因本身也存在于质粒上时，这适用于大肠杆菌，并且底盘基因整合到染色体中，如宿主生物体光杆状菌中的情况。
图10(C)中提供了将高水平的Cre位点特异性重组酶反式包装到大肠杆菌中表达的PVCpnf纳米注射器中的进一步示例。更详细地，发明人构建了实验室大肠杆菌表达菌株，其携带(i)用于例如SEQ ID NO.:93的不对称光杆状菌ATCC43949 PVCpnf操纵子的阿拉伯糖诱导型表达质粒(在Pvc16上具有C-末端FLAG标签，例如紧邻SEQ ID NO.:93的3')和(ii)第二IPTG诱导型表达质粒，其含有Cre重组酶，其具有天然Pnf效应子50个氨基酸前导序列(例如SEQ ID NO.:78的前导序列)的N-末端融合体和C-末端myc标签表位。将PVC操纵子和效应子(Cre+前导序列)共同诱导24小时，并纯化嵌合纳米注射器。蛋白质印迹分析用于确认纯化后FLAG标记的Pvc16帽蛋白(并因此纳米注射器底盘)和反式包装的Myc标记的Cre重组酶的存在。
4.2使用证明了更大的、多样化序列空间的功能性的额外前导序列的反式包装
补充实施例3中概述的数据，图10D证明了使用以下四种另外的前导序列将Cre(反式)包装到(大肠杆菌中的)PVCpnf中(因此证明了更大的序列空间的功能性)：
-泳道1：PAU_02096的前导序列(前导序列＝SEQ ID NO.:71)，实验在图10D中称为“纳米注射器+lopt50::Cre::Myc；
-泳道2：PAK_02075的前导序列(前导序列＝SEQ ID NO.:50)，实验在图10D中称为“纳米注射器+cnf50::cre::Myc；
-泳道3：PAU_02009的前导序列(前导序列＝SEQ ID NO.:68)，实验在图10D中称为“纳米注射器+cif50::cre::Myc；和
-泳道4：PAU_02806的前导序列(前导序列＝SEQ ID NO.:76)，实验在图10D中称为“纳米注射器+gog50::cre::Myc。
这些结果还证明了显示出更大序列多样性的前导序列用于(反式)包装有效载荷的效用。实际上，为了提供进一步的验证，发明人对一组前导序列进行了CLUSTALW序列比较以确定多样性。PVC效应子被鉴定为编码可识别的毒素样结构域的蛋白质，所述毒素样结构域紧接在pvc16结构基因的下游被编码。每个PVC操纵子可以仅编码串联阵列中的单个效应子，或几个不同的效应子基因。系统发生树显示于图10E中，其中本文示例的用于将有效载荷蛋白包装到纳米注射器复合物中的前导序列的身份由不对称光杆状菌ATCC43949PVCpnf操纵子(实线箭头)或发光光杆状菌TT01 PVCunit4操纵子(虚线箭头)或两者来阐述。
从图10E的树可以看出，所示例的前导序列整体分布良好，因此处于或接近最大程度地顺序多样化。
实施例5
尾纤维/结合结构域修饰
已知PVC针状复合物包含尾纤维(参见3D呈现的PVC结构，最右侧图像的最左侧星号)，其被认为允许PVC复合物的细胞类型特异性靶向。发明人已经成功地证明了结合到非天然氨基酸中的尾纤维区域的修饰(例如，野生型序列中的氨基酸取代20种标准氨基酸中的替代氨基酸)不影响尾纤维的表达。
实施例6
证明用前导序列包装的PVC针状复合物将活性(外源)酶/有效载荷物递送到离体
鼠类器官中
概念：获得用于将外源功能性酶递送至哺乳动物组织的数据。发明人已经证明了将称为“Cre”的反式包装的噬菌体衍生的重组酶蛋白递送到离体小鼠胆管类器官中。类器官源自小鼠品系，其中染色体编码的红色荧光蛋白(RFP)报告子的表达通常被侧翼为Cre-重组酶的loxP识别位点的终止信号阻止。如果存在重组酶，则终止信号被重组出来，然后细胞继续表达报告蛋白。这个实验证明背后的一般原理总结在图16A中。
方法：胆管类器官制备：按照Huch等(Regen Med.2013年7月；8(4):385-7.PMID:23826690；DOI:10.2217/rme.13.39)方案，使用“BD扩增培养基”在matrigel中将鼠原代胆管分离并扩增12代成为类器官。然后将细胞以2D接种中并在BD扩增培养基中培养。小鼠基因型:Rosa26基因座中的LSL-Tom报告子+Axin2CreRT(在4OHT处理后可诱导)。将细胞在未包被的聚苯乙烯板中以10,000个细胞/孔的接种密度培养。将纳米注射器制备为在PBS+70％培养基中的30％体积注射器制剂。总体积为100μl/孔。阳性对照代表1:1000(v/v)的500nM 4OHT(在乙醇中)作为重组的阳性对照。阴性对照仅表示1:1000(v/v)乙醇稀释。将细胞接种并生长48h，加入纳米注射器，然后再培养24小时，然后固定(4％PFA RT固定15min)并染色以进行显微镜检查。染色：来自Rockland的一抗抗RFP(1:1000)。二抗为抗兔568(以1:500v/v使用)。在激光共聚焦显微镜上观察样品。
结果：图16B包括来自这些实验的代表性显微照片，证明用加载Cre的PVCpnf纳米注射器处理时，可以在许多细胞中检测到RFP蛋白的信号。由于这些是离体类器官，而不是简单的细胞单层，因此预期给药的细胞数量具有一定的随机性，并且甚至在阳性对照中观察到这一点，阳性对照是小分子诱导物(而不是大的蛋白质复合物)。预期由于这些是类器官，因此将存在一定水平的细胞分化，这可能改变纳米注射器的结合特征。来自该初步运行的另一个有趣的观察结果是，虽然尚未获得关于应用于系统的纳米注射器总量的信息，但发明人证明TAM小分子诱导物似乎不具有比纳米注射器明显更大的组织穿透，表明它们的分布能力不主要是受它们尺寸的阻碍。
其他解释：总之，发明人已经证明了将外源酶递送(例如给药)至细胞靶标的能力。此外，通过证明提供导致转化细胞的DNA改变的能力，该“纳米注射器+Cre”实验是有前景的生物技术工具/辅助概念的证明。因此，该实验证明了外源有效载荷(病毒而不是细菌的蛋白质)，特别是核酸修饰酶的用途。显然，Cre酶以功能性方式递送，并且能够穿过细胞内部到达核以影响其DNA修饰改变。
实施例7
将MAD7位点特异性重组酶(外源有效载荷)反式包装到大肠杆菌中表达的PVCpnf
纳米注射器中
概念：与(实施例6的)Cre数据和本文提供的包装的有效载荷的其他实例一样，发明人已经证明了通过前导序列将Cas-样酶MAD7包装到纳米注射器中。这是本文所述的有效载荷的最大外源实例(MAD7＝147.9kDa)。
方法：简而言之，在大肠杆菌中同时表达(在诱导后)底盘基因和MAD7基因(后者用C-末端Myc标签标记用于检测，以及本文所述的用于纳米注射器结合的前导序列)。在收获和纯化纳米注射器复合物后，通过斑点印迹分析探测有效载荷包装(例如，用于检测Myc标签)。本文所述的纯化方法(使用超速离心)可用于选择(例如过度)高分子量蛋白质复合物/生物物质，使得能够回收纳米注射器和它们携带的任何货物(有效载荷)。“松散的”/未包装的有效载荷保留在溶液中并且不经受足够的离心力，因此在纯化期间损失，除非包含在大得多的纳米注射器“壳”内(即，成功包装时)。图17证明了MAD7的成功包装。
实施例8
将凋亡诱导有效载荷反式包装到在大肠杆菌中表达的PVC pnf中
使用图10C中描述的大肠杆菌PVCpnf前导序列::有效载荷::Myc反式包装系统(PVCpnf前导序列＝SEQ ID NO.:78)，发明人证明了反式包装至少两种促凋亡人源担保之序列或肽(例如SEQ ID NO.:109和SEQ ID NO.:111的序列)的能力。将Pnf效应子蛋白前导序列(例如SEQ ID NO.:78)与N-末端融合，并且将Myc表位标签与C-末端融合。蛋白质斑点印迹分析(类似于实施例7)证实了这些人源蛋白质在纯化的纳米注射器中的存在(图18)。
实施例9
通过纳米注射器递送(反式)包装的促凋亡人多肽在培养的离体人细胞中诱导凋
亡的证明
初步测试已经证实了使用在大肠杆菌中产生的PVCpnf纳米注射器递送反式包装的人蛋白质序列(例如根据实施例8包装的)并诱导来自人供体的离体循环PBMC细胞凋亡的能力。该测定是来自仅暴露于包装的纳米注射器20分钟的细胞的TUNEL染色显微镜分析。结果显示在图19A中，证明(通过成功诱导凋亡)tBid p15片段和BaxBH3结构域的递送。
·tBid p15片段(SEQ ID NO:109)是正常人细胞凋亡调节途径的一部分。细胞效应：Bcl-2家族的促凋亡成员。Bid的C-末端部分(tBid)易位至线粒体，在那里它诱导细胞色素c的释放。Bid通常被胱天蛋白酶8从其潜在的胞质全长pro-Bid形式切割。
·BaxBH3(aa59-73)(SEQ ID NO:111)是最小BH3结构域合成肽，其包含限定的BaxBH3结构域的关键的15个残基。细胞效应：这15个残基含有足够的信息以结合并功能性地拮抗Bcl-xL并特异性地诱导Bax/Bak。似乎消除了Bak/Bcl-2相互作用-释放促凋亡因子。
现在描述更详细的将促凋亡人肽递送到离体外周血单核细胞(PBMC)中的测试。本研究的目的是研究负载促凋亡肽的PVC纳米注射器是否可以在离体人外周血单核细胞中诱导凋亡。首先使用台盼蓝染料排除测定评估纳米注射器的任何即时细胞毒性，然后通过使用TUNEL测定评估凋亡反应。
细胞生存力的台盼蓝排除试验：台盼蓝是通常用于选择性着色死组织或细胞的重氮染料，因此，死细胞在显微镜下显示为独特的蓝色，而具有完整细胞膜的活细胞或组织保持无色。由于活细胞被排除在染色之外，因此该染色方法也被描述为染料排除方法。台盼蓝通常用于评估组织或细胞生存力。将合适数量的细胞(2×105)暴露于纳米注射器和空纳米注射器20分钟。将合适体积的细胞(30μL)加入等体积的0.4％台盼蓝中，并使用血细胞计数器计数存活(未染色)和死亡(染色)细胞的数量。在3个浓度下测试每种化合物。针对每种化合物在每种浓度下测试来自两个独立人类供体的血细胞，并且每种样品一式两份测试。
用于显微镜检查的细胞的处理和制备：在2个独立测试中，在3个测试浓度下，用两个嵌合纳米注射器(例如，负载有外源促凋亡肽)处理20分钟后，测定来自两个独立健康人供体的外周血单核细胞(PBMC)的生存力。通过离心收获PBMC，并以1×106个细胞/ml重悬于培养基中。将细胞在2.5％福尔马林中固定并在室温下孵育20min。通过用70％乙醇喷雾并使其风干来制备聚-L-赖氨酸包被的载玻片。将细胞离心30秒。除去上清液，将细胞重悬于200ml dH2O中。向每个载玻片/固定中加入5ml细胞悬浮液。每个载玻片进行两次固定以允许一式两份进行染色。使细胞悬浮液风干。
PBMC细胞生存力测定的结果：台盼蓝活力测定证实了PVC制剂本身对取自健康人供体的PBMC没有立即毒性(表2)。纳米注射器处理显示>60％活力，表明在最大剂量浓度下的低毒性(表2)。然后，发明人继续测试嵌合纳米注射器诱导细胞凋亡的能力。
表2.在3个测试浓度(v/v稀释)下暴露于每种化合物20分钟后，来自两个独立人类血液供体的外周血单核细胞(PBMC)的生存力。PBMC对照未处理。
使用TUNEL测定测试嵌合纳米注射器诱导的细胞凋亡：然后使用TUNEL测定来鉴定固定在载玻片上的单细胞悬浮液中的凋亡细胞核。在该测定中，末端脱氧核苷酸转移酶(TdT)与响应凋亡信号因子而产生的DNA片段暴露的3'-OH端结合。这又催化生物素标记的脱氧核苷酸的添加，其可以使用链霉抗生物素蛋白-辣根过氧化物(HRP)缀合物来检测。二氨基联苯胺(DAB)与HRP标记的样品反应，在DNA片段化位点产生不溶性棕色底物。甲基绿复染能够使正常细胞和凋亡细胞可视化。
测定了人PMBC暴露于纳米注射器后凋亡的诱导。TUNEL测定试剂盒(Abcam)用于检测凋亡细胞。按照制造商的说明书进行测定。简而言之，用100μL蛋白酶K溶液覆盖载玻片5分钟，用1×TRIS缓冲盐水(TBS)冲洗载玻片。纳米注射器或DNA酶I阳性试剂盒对照的处理在室温下进行20分钟。用TBS冲洗载玻片。然后将载玻片与TdT平衡缓冲液一起孵育30分钟，然后加入TdT标记反应混合物。将载玻片在37℃下孵育19分钟。然后用TBS洗涤载玻片，然后施加终止缓冲液并在室温下孵育5分钟。再次用TBS洗涤载玻片，然后在室温下加入封闭缓冲液10分钟。通过将缀合物应用于样品30分钟来进行检测。用TBS冲洗载玻片，然后施加DAB溶液15分钟。用dH2O冲洗载玻片，然后用甲基绿复染。将载玻片在100％乙醇中脱水，然后在二甲苯中脱水，并用玻璃盖玻片固定。所有染色一式两份进行。指示细胞凋亡检测测定中阳性染色的凋亡终点由深棕色(DAB)信号表示。较浅的棕色阴影和/或蓝色/绿色至绿色/棕色阴影表示凋亡的非反应性阴性细胞。
通过在载玻片上选择5个随机细胞切片进行分析，计数阳性染色细胞(深棕色)和阴性染色细胞(蓝色或浅棕色)，并确定显示凋亡小体的细胞的百分比。
为了产生阳性对照，在下文详述的蛋白酶K处理步骤之后，在室温下用1μg/ml DNA酶I(试剂盒阳性对照)处理载玻片20分钟。DNA酶I处理使正常细胞中的DNA片段化以产生与凋亡期间产生的那些相同的游离3'OH基团。通过在处理阶段期间在反应混合物中用dH2O替代DNA酶I来产生阴性对照。
PMBC细胞凋亡测定的结果：在用完整的tBID和Bax加载的纳米注射器处理后，用适当的阳性和阴性试剂盒对照，进行使用PBMC的TUNEL染色。进行处理20min以确定纳米注射器是否引发凋亡信号。包括阳性对照(DNA酶I处理)和阴性对照(无DNA酶I处理)。结果显示含有tBID或Bax的两种纳米注射器在PBMC上显示强凋亡信号(分别为89％和78％阳性)。阳性对照显示了强凋亡信号(79％)，而阴性对照未显示凋亡信号(100％阴性)。还观察到纳米注射器处理的样品中附着细胞数量的显著损失，推测表明快速和全面的细胞凋亡反应，并且在洗涤后不能保留。注意到这种效果比试剂盒阳性对照甚至更明显，表明反应更快。代表性显微照片显示在图19B中。
结论：推断负载tBID和Bax的纳米注射器能够快速诱导人外周血单核细胞的广泛凋亡。此外，台盼蓝染料排除测定已经证实了这些嵌合纳米注射器不会对细胞造成快速致死裂解或广泛的膜损伤。
实施例10
前导序列和PVC针状复合物的实际效用的示例-非典型(非光杆状菌属)有效载荷
的胞内递送
(1)抗MDM(p53抑制剂)抗体与本文所述的前导序列连接，并与PVC针状复合物一起表达以包装在其中。将分离的PVC针状复合物(包含抗体有效载荷)与肿瘤接触以用于抗体的胞内递送(所述肿瘤细胞的特征在于具有用于MDM抑制的p53活性的高MDM抑制)。肿瘤被抗MDM抗体的活性抑制。
(2)PVC针状复合物用于(胞内)递送抗肿瘤肽疫苗以激活MHC-I依赖性细胞毒性T细胞淋巴细胞(CTL)应答。递送酪氨酸酶相关蛋白2(TRP2)肽疫苗以增强对CTL的交叉呈递发生和针对表达TRP2的肿瘤的抗肿瘤作用。肿瘤被肽疫苗的活性抑制。
(3)PVC针状复合物用于(胞内)将核因子-kB抑制剂(其用于控制炎性病症，如类风湿性关节炎)递送至细胞。该细胞随后证明了促炎细胞因子的表达降低。
(4)PVC针状复合物用于(胞内)递送T3SS有效载荷(其抑制NF-κB和MAPK途径)。这用分离的(纯化的)PVC针状复合物完成，而不需要PVC针状复合物保持与其来源的细菌细胞相缔合。
(5)PVC针状复合物用于(胞内)向细胞递送抗凋亡肽，包括BH4、Bcl-xL-蛋白和/或c-Jun N-末端激酶的肽抑制剂(其可以保护心脏和脑免受缺血性损伤(对组织的血液供应的限制，导致细胞代谢所需的氧和葡萄糖短缺))。例如，通过JUN激酶的20个氨基酸结合基序的JUN激酶抑制是足够的。例如细胞色素c在细胞中的释放被抑制。
(6)PVC针状复合物用于(胞内)递送烟酰胺腺嘌呤二核苷酸醌内部氧化还原酶(Ndi1)，复合物I的单亚基酵母类似物(其提供显著的心脏保护作用)至复合物I缺陷型突变细胞。Ndi1蛋白正确地靶向线粒体内膜的基质侧，并将NADH氧化酶活性恢复到复合物I缺陷型细胞。
(7)PVC针状复合物用于将PHOX复合物(其用于酶替代疗法以恢复慢性肉芽肿病中ROS的产生)的两个必需亚基之一递送至慢性肉芽肿病细胞。观察到ROS产生的恢复。
(8)PVC针状复合物用于(胞内)递送(例如肌内)肌管蛋白(其用于改善X连锁肌管肌病患者的局部和远处肌肉性能)。观察到磷脂酰肌醇3-磷酸和磷脂酰肌醇(3,5)-二磷酸的肌管蛋白-脱磷酸化。
(9)PVC针状复合物用于(胞内)将重组酶“Cre”(其能够切除限定的遗传盒)递送到小鼠细胞系中，其中基因组具有侧接mCherry基因上游的终止信号的loxP重组位点。Cre有效载荷切除重组位点，并去除终止信号，允许mCherry基因在细胞中表达。
(10)PVC针状复合物用于(胞内)递送对胞内组分具有亲和力的～15kDa纳米抗体(抗体片段)。检测纳米抗体-胞内复合物。
(11)PVC针状复合物用于胞内递送(例如递送到昆虫细胞中)用于昆虫作物害虫和动物寄生虫的非典型(非光杆状菌属)多肽毒素。观察害虫的抑制。
(12)PVC针状复合物用于(胞内)将核酸酶(例如Cas9和/或Mad7)递送到包含指导RNA的靶细胞中。核酸酶进行定点基因失活。
上述说明书中提及的所有出版物均通过引用并入本文中。在不脱离本发明的范围和精神的情况下，本发明的所述方法和系统的各种修改和变化对于本领域技术人员将是显而易见的。尽管已经结合具体的优选实施方案描述了本发明，但是应当理解，所要求保护的发明不应不适当地限于这些具体实施方案。实际上，对于生物化学和生物技术或相关领域的技术人员显而易见的用于实施本发明的所述方式的各种修改旨在落入所附权利要求的范围内。
序列
在任何以下SEQ ID NO中指示初始Met氨基酸残基或相应的初始密码子的情况中，所述残基/密码子可以是任选的。
SEQ
ID
NO:1(PAK_1985)
MMREYSNEDDFIKEKTNLVKSENVEADNYLETEYLTYLAKLIGMTERENHHLNSIKLIDDIIELHNDRKGNKLLWNDNWQDKIIDRDLQSIFKKIDEMVSEFGGLEAYKDIVGENPYDPTEPVCGYSAQNIFKLMTEGEYAVDPVKMAKTGKINGNQFAEKLEHLNSSNNYVALINDHRLGHMFLVDIPSTNRERVGYIYQSDLGDGALPALKIADWLKSRGKESINVNKLKKFLNDEFTMLPDNEQKGLIAEIFDLNKDIDSVKSGKIKKDKAVDIYLREYDINDFISNIEKLKTKLA
SEQ
ID
NO:2(PAK_1987)
MFQNRIRNEKTTQSGKGKTLDRMTDSLYLEIPNVEAVTLAYQKLTSKYRKFDNKTKLILDSSDEFSQLKSEKQRKGFSKSGLKNNGVSDRKFIYTKNALKNFAAHAGYEHNGHYEDEFVNFKDNNKNLAKGKLFPGISLIERRKLSIVKNKEGKWEHKETDEAEAYKVTDIEKFISGVRSMYLQGNTFLHAKTEALIRKHIANNENILPTMAGIAGLHAEVQALNNLFISGDKGTKKREKWKYIRNMLESSIFTQRLTTGQAGKDFAACHNCSGILSSPVNVITGKVESAGDNFLSTLSRYKTSQESPI
SEQ
ID
NO:3(PAK_1988)
MEREYSEKQKNPSKLSRKTAISERIAALERSGLSNSNQPVPQFARPYTSNRPVVNINPGRSSIAVATANSTSPVNIPTPAPASPDKLLPSTSCDTTSSILIVGKYNLELTSQGKIVVFRGDNRTPEQIVAAGGFYPWSKQDVGKIKKELIDEFIEIGPSAHMMGHVRSPNKNYVSTGMNMDSGGFGEQSNYLYKMEIPGLKPQDMNERTLGEKIRQDKRGINYPHFLMSHLTLAESEFVAMIPARSEELTFITPIPLSYITSYRKRGTNTWLPMPLKK
SEQ
ID
NO:4(PAK_2075)
MSNYEYDIVTQHDTYQIKDNEYTVVNGKYWQYEQEGNKNNNKVSISLMKENQNDPVWITSDIKEISLYIIENLFSYHKFSAELQHTLKNAVKAVFNEYSEIKYSELLHNINNIFNLFFIKIYNTSDIDTAINILTAKIEIYDKLEKINQDKTDSNNTNVDIWEELGINAEEPLLKIYRQAFSTGDIDDEVYSDALLTFMSDGNLELGDKEKSDYNQRIKDKTDLFESYKKGIEKVASLITTNNINPGIPITYPETEKSINIGDDLLLAQLAKEEIALKKQNRTEYSQQDIFELQTLQAAKYHLLILSSLGALLYQIAPNVEKMTKGHGDYRDIIFSQEQAESLFKKHNIQYDTNHVLSQESKHIEMEGCIILTAAIIYRMRKENATVEQALNYSTLETIKLFENDKKKLNPFNTNNVKPAGYFSFIDFKKRDKFDSQYNFNEQFNVYKNKYSHYESISFSKLILSSPAAQLTAEEIVNPPEEAFLYSVEQGMGNVAMIKMYQGNWLVISTIQGGVKAKKYSRQQVDSNPTLRAMSKPNALFLIERKMETGMGILMPNMMVNTGKRLFPTGYERAKTLSGFAETSRYKNSYNAFWNDYYGITSGMNVGISFTGSPKFNFYKEENLLSVTATIIQQGLNDIAIKSKQALDITSGWHIAATILIPFYNVIYKSTTDSEYELTGEDIGSIVFDTANVLLVVATLGMSLTESMAAKVTQTTLRLRQAGLTGRALITAVVRTLPEHGIITLRQSSGIILGGLIDLIEPLPIRSTLTLTYRGVINAVGAMRNSIKLEKSFADIFGKSTRGLGKLKNEWKVSNLPLEEIVPHSNGGEIYKGIYSIRPTNPETAVKQNFYIKEAGANYQVKWDDANHTWRVVNPTYPEQFSYWPAVKLDKNGHWVTHADVSNKFLILEQSKRIDQELEAAHSNINNDNILDAFIHINTAFKDCERYDIDKLSDITDTLTHFFEKSLKPGDKKAIFSTEIMSIQQAWIREVILPLQNNSSISIEKINAIKTELPYLLRKTFPIESQLPNQLVANKIALAIEEIPNTRIPKYTSGNISKTVQYTSLLENNHVDIPPVGITITGNDTFINQVTRVLSEIDEIPSGNIVIQELEKQGLNIQPPTMNDIVREKNGQFYANNSAGSHIAFDPENHLIGTEEKLIDEPWRTREPAIALYHEMLHIYYNRYPTWFTSIDNKVIDQKVSGGFSLLEESRIVGTKYYVNDKNTLFDFNDSDYLLENNSALLTENRFRAEYAIFKNKSEYVIRPYSGKGDSQIPLTKTKININESHRNVMGVGSGKPEKMPNESATDYRNRVREWRKANKQPEADIGTGDMRKTKAEARVKLLKENYPQFEPQKIELGGAFQLWTVPNEPANKLMLSSHGYFFSDSAATQVPAGKTIQFLGPHGKTLLEAPENPLYSPFDVTLGNSGFTVQPYATIESGNKAGLGSVKIGDKTFTVNDIQNIATDDVENYLLATGVEANASNHGKVRNYGIKYYEKMPDEEVKAAIWKNRADETSTHKYDALLVSPEAGNRKKLSDIFALMKTDERMSKYDEITFVACREELNRINMKSIHDTGLGGGYEPKLEPTVILSRRRREATFTADGAIIYSIIAVNLHHNFITEEIVGIAPFLFINN
SEQ ID NO:5(PAK_2077)
MEHEYNEKEKQRNSAIKLNDAIRNNEENMDMTSPLELNFQNTNRKSRGLRERFSATLQRNLPGHSMLDRELTTDGQKNQESRFSPGMIMDRLMHFGVRTRLGKVRNSASKYGGQVTFKFAQTKGTFLDQIMKHKDTSGGVCESISAHWISAHAKGDSIFNQLYVGGKKGKFHIDTLFSIKQLQMDGYLDDEQSTMTEYWLGTQGMQPNIQRNDDTDEHSSKVVGETGNRGTKDLLHAILDTGDKGSGYKKISFLGKMAGHTVAAYVDDQKGVTFFDPNFGEFSFPDKTSFSHWFTDDFWPKSWYSLEIGLGQEFEVFNYAPEAP
SEQ
ID
NO:6(PAK_2892)
MPNKKYSENTHQGKKPLMKSEANNEHDIQNSSLGIGLDLNSMMGNSSTSLSHIQDYSFWKENISEYYKWMVVVKAHLKQLDWTLKSMDSPESAGTNIAKNTGTTALQTLLNTGGSIAGAAIGGAIGSAIAPGVGTIAGMGIGALAGTGLNYLNDTVIEKLNEKLEIAYPYPKTRNMIFDINNYDKNPIIKAIKKKTNKDNLKVTAGSSLTSQLVGKVTSPIKFPAYKLADLAIALAGLSSDKARHILDFTDSIREVLNESHSDAVAFMRKNYGDNAMGLAGLSSRIK
SEQ ID NO:7(PAK_2893)
MEREYSEKEKHKKRPIQLRNSIEQHEEETANNSLGLGLDLNQATNPPKVPKDNYNEENGDLFYGLANQRGRYIKSVNPNFDPDKINSSPMIIDVYNNNVSNTILNKYPLDKLVKLSGNPQKYANNIKVENSLQQDVASSKRGWYPLWNDYFKTGNENKKFNIADIYKETRNQYGSDYYHTWHTPTGAAPKLLWKRGSKLGIEMAASNEKTKIHFVLDGLNIQEVVNKQKGSTPLEQGRGESITASELRYAYRNRERLAGKIHFYENDQETVAPWEKSPELWQNYIPKNKNQNESSTPQRNNGTLYRLGGPFRKLRASLRKRS
SEQ
ID
NO:8(PAK_2894)
MMEHEYSKEEEKKRQQSKPNNATHDESNLPLELEKHFNARTPATAHSKWFTYENDTEVELTTERIKEIFSNKQPKIIIAGDGHNKPPFQYAKNIPDVNSSFDAGTLQLYIEATDEQINENNPEYIPKEFMAKPGLFTNKNRRAEIVGWEDSELSNAMKEMFELSDKSTREKLTPEETSSFYKLHETAIRHFFRPEFNQLRDEFFEILAKAGSNRELDKIALEMIGFTSGTWRDEYINPTLAEKIAKHAAEKENHTFVVSIGDAHLSENPMQEYLNKRRNGGEFKHQIIFTRDKRPILPDNMKTGNKNS
SEQ
ID
NO:9(PAK_3525)
MLKYANPQAVPTQRTKNTAKKPSSSSSFDGQLELSNGEWSKHSEMGLKRGGLINSIRRRIARNGNIGRFNELIDSEAKKWPSEPVDKNIHMIWIGTRNISEKNIKLSIDTAKKNPDYNTSIIYDSGISGHEGARNFMLEKFEGSNVNXSLAFPKGIGVMREYAPEAGKATAFPNTPIAVTKNNPIINKTLDLAVGNYQRGEKNVLKLAGPDVFTQALYQEIPGLNSKVLNAQLDQFELAKRQALGLPLEKPKSFADEKLTSVEKEKINRPYQSMRGLSGHVMNGADHSWAVDTEVLGH
SEQ
ID
NO:10(PAT_00148)
MMREYSNEDDCTKEKTNLVKSENVEADNYLEMEHLTYLAKLISMTERENHHLNSIKLIDDIIELHNDRKGNKLLWNDNWQDKIIDRDLQSIFKKIDEMVSEFGGLEAYKDIVGESPYDPTEPVCGYSAQNIFKLMTEGEYAVDPVKMAKTGKINGNQFAEKLEHLNSSNNYVALINDHRLGHMFLVDIPSTNRERVGYIYQSDLGDGALPALKIADWLKSRGKESINVNKLKKFLNDEFTMLPENEQKGLIAEIFDLNKDIDSVKSGKIKKDKAVDIYLREYDINDFISNVEKLKTKLA
SEQ ID NO:11(PAT_00149)MIFKMLNLAVFYLLGNIFHYLICQKFICYFCSVLKSVTMFLTKVAVQIALYLNILPTMAGIAGLHAEVQALNNLFISGDRGTEKRENWKYIRNMLESTIFTQRLTAGQAGKDFAACHNCSGILSSPVNVITGKVESAGGNFFINIISI
SEQ
ID
NO:12(PAT_00150)
MEREYSEKPKNLSQLSRKTAISERRAMFERNASSNNEQPVPQFARSYTSNRSVVNINPGRSSIAVVTANSTSPVNISTPAAASPDKLLPSTSCDTTSSTLTVGKYKLELTSQGKVVVFRGDNRTPEQIVAAGGFGEQSNYLYKMEIPGLKPQDMNERTLGEKIRQDSRGN
SEQ
ID
NO:13(PAT_00152)
MKYDPRLRTWVEDDFDYEKNFKKQTDYINYKDLEKQLKENVDYYALLDENEAIIFLKELGCDIKSFLNDTAFPVTDVLSNFAGNIKDALGVFKVAKNFKPINIGIFTYIINELKGKGIKAIEYLGKNGERYIKLTDRPGIRKYLNATRYLINNKKIMEVGIGSVAMEGSIVKGARFGVIYSAAYRSVELMFKSEYDLTNFFVNLSMDMAKIIVATIIAKSTVAAATSFVVTAALSTTAIAIGVFIIGALVVWGLMWLDDEFKISETIIRRLKEHKVKTPISTYHSDQIFNAWGRYYRG
SEQ ID NO:14(PAT_02308)
MPNKKHSENTHQGRKPLIKSEANNEHDIENSSLGIGLDLNSTIGNNSASLSQIQDYSFWKENISEYYKWMVVVKAHLKQLDWTLKSMDSSESAGTNIAKNIGTTALQTLLNTGGSIAGGAIGGAIGSAIAPGVGTIAGMGIGALAGTGLNYLNDTVIEKLNEKLEIAYPYPKTRNMIFDINNYDKNPIIKAIKKKTNKDNLKVTAGSSLTSQLVGKVTSPIKFPAYKLSDLAISHNRALAGLSSDKARHILDFTDSIREVLNESHSDAVAFMRKNYGDNAMGLSGLSSRIKGEKLTLATLARTRNKIENRINSINKQTLKLSSKNSNE
SEQ
ID
NO:15(PAT_02309)
MEREYSEKEKHKKRPIQLRNSIEQHEEETANNSLGLGLDLNQATNPPKVPKDNYNEENGDLFYGLATQRGRYIKSVNPNFDPDKINSSPMIIDVYNNNVSNTILNKYPLDKLVKLSGNPQKYANNIKVENNLQQDVASSKRGWYPLWNDYFKIGNENKKFNIADIYKETRNQYGSDYYHTWHTPTGAAPKLLWKRGSKLGIEMAASNEKTKIHFVLDGLNIQEVVNKQKGSTPLEQGRGESITASELRYAYRNRERLAGKIHFYENDQETVAPWEKSPELWQNYIPKNKNQNESSTPQRNNGALYRLGGPFRKLRASLRKRS
SEQ
ID
NO:16(PAT_02310)
MMEHEYSKEEEKKRQQSKPNNATHDESNLPLELEKHSNARTSATAYSKWFTYENDMEVELTTERVREIFSNKQPKIIIAGDGHNKPPFQYTKNIPDVNSSFDAGTLQLYIEATDEQINENNPEYIPKEFMAKPGLFTNKNRRAEIVGWEDSELSNAMKEMFELSDKSTREKLTPEETSSFYKLHETAIRHFFRPEFNQLRDEFFEILAKAGSNRELDKIALEMIGFTSGTWRDEYINPTLAEKIAKHAAEKENHTFVVSIGDAHLSENPMQEYLNKRRNGGEFKHQIIFTRDKRPILPDNMKTGKKNS
SEQ
ID
NO:17(PAT_02956)
MSNYEYDIVTQHDTYQIKDNEYTVVNGKYWQYEQEGNKNNNKISISLMKDNQNDPVWITSDIKEISLYIIENLFSYHKFSAELQHTLKNAVKAVFNEYSEIKYSELLHNINNIFNLFFIKTYNTSDINTAINILTAKIEIYDKLEKINQDKTDLNNTKVDIWEELGINAEEPLLKIYRQAFSTGDIDDEVYSDALLTFMSDGNLKLGDKEKSDYNQRIKDKTDLFESYKKGIEKVASLITTNNINPGIPITYPETEKSINIGDDLLLAQLAKEEIALKKQNRTEYSQQDIFELQTLQAAKYHLLILSSLGALLYQIAPNVEKMTKGHGDYRDIIFSQEQAESLFKKHNIQYDTNHVLSQESKHIEMEGCIILTAAIIYRMRKENATVEQALNYSTLETIKLFENDKKKLNPFNTNNVKPAGYFSFIDFKKRDKFDSQYNFNEQFNVYKNKYSHYESISFSKLILSSPAAQLTAEEIVNPPEETFLYSVEQGMGNVAMIKMYQGNWLVVSTIQGGVKARKYSQQQVDSQPTLRAMSRPNALFLIERKIMIGIGIFMENQIVNTGKRLFPTGYERAKTLSGFAETSRYKNSYNAFWNDYYGITSGMNVGISFTGSPKFNFYKEENLLSVTATIIQQGLNDIAIKSKQALDITSGWHIAATILIPFYNVIYKSTTDSEYELTGEDIGSIVFDTANVLLVVATLGMSLTESMAAKVTQTTLRLRQAGLTGRALITAVVRTLPEHGIITLRQSSGIILGGLIDLIEPLPIRSTLTLTYRGVISAVGAMRNSIKLEKSFADIFGKSTRGLGKLKHEWKVSNLPLEEIVPHSNGGEIYKGIYSIRHTNPETAVKQNFYIKEAGANYQVKWDDANHTWRVVNPTYPEQFSYWPAVKLDKNGHWVTHADISNKFLILEKSKRIDQELEAAHSNINNDNILDAFIHINTAFKDCERYDIDKLSDITDTLTHFFEKSLKPGDKKAIFSTEIMSIQQAWIREVILPLQNNSSISIEKINAIKTELPYLLRKTFPIESQLPNQLVANKIALAIEEIPNTRIPKYTSGNISKTVQYTSLLENNHVDIPPVGITITGNDTFINQVTRVLSEIDEIPSGNIVIQELEKQGLNIQPPTMNDIVREKNGQFYANNSAGSHIAFDPENHLIGTEEKLIDEPWRTREPAIALYHEMLHIYYNRYPTWFTSIDNKVIDQKVSGGFSLLEESRIVGTKYYVNDKDTLFDFNDSDYLLENNSALLTENRFRAEYAIFKNKSEYVIRPYSGKGDSQIPLTKTKININESHRNVMGVGSGKPEKMPNESATDYRNRVREWRKANKQPEADIGTGDMRKTKAEARVKLLKENYPQFEPQKIELGGAFQLWTVPNEPANKLMLSSHGYFFSDSAATQVPAGKTIQFLGPHGKTLLEAPENPLNSPFDVTLGNSGFTVQPYATIESGNKAGLGSVKIGDKTFTVNDIQNIATDDVENYLLATGVEANASNHGKVRNYGIKYYEKMPDEEVKAAIWKNRADETSTHKYDALLVSPEAGNRKKLSDIFALMKTDERMSKYDEITFVACREELNRINMKSIHDTGLGGGYEPKLEPTVILSRRRREATFTADGAIIYSIIAVNLHHNFITEEIVGIAPFLFIDN
SEQ
ID
NO:18(PAT_02957)
MEHEYNEKEKQRNSAIKLNDAIRNNEENMDMTSPLELNSQNTNRKSRGLRERFSATLQRNLPGHSMLDRELTTDGQKNQESRFSPGMIMDRLMHFGVRTRLGKVRNSASKYGGQVTFKFAQTKGTFLDQIMKHKDTSGGVCESISAHWISAHAKGDSIFNQLYVGGQKGKFHIDTLFSIKQLQMDGYLDDEQSTMTEYWLGTQGMQPNIQRNDDTDEHSSKVVGETGTKGTKDLLHAILDTGDKGSGYKKISFLGKMAGHTVAAYVDDQKGVTFFDPNFGEFSFPDKTSFSHWFTDDFWPKSWYSLEIGLGQEFEVFNYAPKEP
SEQ ID NO:19(PAT_03171)
MFKYDTSEKMAKFGKGKTSDGMLLDTLYLEIPDEKAVMSAYKSQILDELRNFSEKTHSFFSGKKPLYSKKYLANLAAHAGYVHVTDYNSIGNYKDGFVNFKDNSRNLAEGKLFPGIRLIKRPKLSIVRDKETERWKKQESDEADAYEITDIESFISGVRDMYSRANVDLHPVIESLIRNHIVNNDHVLPTMAGIAGLHAEVQALNNLLILADGRAGKIVGGRKIEEYMQDMLKSFIFTQRLTTKQAGNDFAACHNCSGILSVPANVITGKVASAGSNFSLILSRYKNSQESPI
SEQ
ID
NO:20(PAT_03172)
MLKHANPQTVSTQRTKSTAKKPSSSSSFDRQFELSNSENQPGEGNKDWTIKGWRQRFADRSLNKGHISPLMNKGLLVGSEEALINVPVVAHRYDSSHQLTDAGPLKADSHSNNLDPFYGVVTGFRGDQVTSSESGSGSIGGHWGKNTLDSNITGINVVNGASGTVGIRIALKDIQHGAPVIVTSGALSGCTMVYAVKNGYFFAYHTGQKPGDKEWKTGRQGVVATYRSHQALSPDSEPMAVGEQNNDLVNIFASYDQGIITYMGKPGVIIDNTAENVGVFNYDEVKLEKPDIRAGYSYALLAKDDKGKVNVKVLSEDVIVPLGNKGKTIKAINSLKKRLL
SEQ
ID
NO:21(PAT_03177)
MPRYANYQINPKQNTKNSHGKSSSSNFSSGYFSSSNNSLDDSLIRQQVKREFIWEGHMKEIEEASRLGNFAVSFRAAGGPTLRALGKGAAAKGHDILEKTIKPGSINKAYPKDEASNVIKKVQEAGIEGYVGHWDKKTGRLLGIYMSSGHGLSDEQVNGKIYPIDLNNLEASLSALKTKENWAALPFTGDYDMHDMISFTGQPHSVPSNSSEERKIIDRINRLVARSDPNRPFGDIEHNVIRHGAQVSYPAFAMDKEKEEIKKHGGIVKAVAEPGEFPVAIVSKGKWTIANNIDELNQFYNSIGAKMKVSWKPGAENPGFVSNPQRPGMARFSRKR
SEQ
ID
NO:22(PAU_02009)
MMREYSKEDDCVKEKTNLAESENVEADNYLEMDCLNYLAKLNGMPERKDHSLNSTKLIDDIIKLHNDRKGNKLLWNDNWQDKIIDRDLESIFKKIDEMVSEFGGIEIYKDIVGENPYDPTEPVCGYSAQNIFKLMTEGEHAVDPVKMAQTGKINGNEFAEKLEQLNSSNNYVALINDHRLGHMFLVDIPSTNREKVGYIYQSDLGDGALPALKIADWLKSRGKESINVNKLKKFLSNEFTMLSESEQKELIAEIFDINKDIANVKLGKIKKDKAVDVYLREYDLNDFISNIEKLKTKLV
SEQ
ID
NO:23(PAU_02010)
MPIIGHKEDLIRTERSSVDLTRSSNNRQTDNLELNIPQHKRDNKDIEHAVIYGFSQHRGPEMQKAFADNKNPVTIDEYNAGLGIMGELSLSDYFRISQDLKENRLPELNEKNIQNHSLKYFDAMGVNMKSADPNVKEEAKEQQRAYTRSWGFYMMENKEKLDIQSKINNLIPKKKSFFSKSPGEDEYKKLDEFILKNSNGSNLTIPKQRKILMKFASAKNAVDVTKNLSGEEQTWLKDIIATAFFRQTSKLGMSWFIEQLASPDFRFVIVGFNGEELTTDQIRSNKPWKHGNRRKEGASEYAEPITFSEIRHAHRKGYDSKINFIKK
SEQ ID NO:24(PAU_02095)
MISTFDPAICAGTPTVTVLDNRNLTVREIVFHRAKAGGDTDTLITRHQYDLRGNLTQSLDPRLYDLMQKDNTVQPNFYWQHDLLGRVLHTVSIDAGGTVTLSDIEDRPALNVNAMGVVKTWQYEANSLPGRLLSVSEQSANEAVPRVIEHFIWAGNSQAEKDLNLAGQYMRHYDTAGLDQLNSLSLTGAHLSQSLQLLKDDQMPDWAGDNESVWQNKLKNEVHTTQSTTDATGAPLTQTDAKENMQRLAYNVTGQLKSSWLTLNGQLEQIIVKSLAYSESGQKIREEHGNGVVTKYSYEPDTQRLINITTQRSKGHVFSEKLLQDLLYEYDPVGNIVSILNRAEATHFWRNQKVSPRNTYTYDSLYQLIQSTGREMADIGQQNNKMPTPLVPLSSDDKVYTTYTRTYSYDRGNNLTKIQHRAPASHNIYTTEITVSNRSNRAVLSHNGLTPREVDAQFDASGHQISLPTGQNLSWNQRGELQQATTINRDNSATDREWYRYNAGSARILKVSEQQTGNSTQQQQVTYLPGLELRTTKSGTNTTEDLQVITMVETERTQVRILHWSAGKPNDIANNQVRYSYDNLIESNVMELDTKGKIISQEEYYPYGGTAIWTARNQIEASYKTVRYSGKERDKTGLYYYRHRYYQPWLGRWLSADPAGTVDGLNLYRMVKNNPIRYQDESGTNANDKAQAIFKEGKKIAINQLKIASNFLKDSKNSENALEIYRIFFGGHQDIEQLPQWKKRIDSVIYGLDKLKTTKHVHYQQDKSGSSSTVADLNVDEYKKWSEGNKSIYVNVYADALKRVYEDPLLGREHVAHIAIHELSHGVLRTQDHKYIGVLSSPGSHDLTDLLSILMPPANEQDRTEKQRRATGARKALENADSFTLSARYLYYTAQDPNFLSSLRKAHRDFNNKKTDRLIIRPPERR
SEQ
ID
NO:25(PAU_02096)
MEREYNKKEKQKKSAIKLDDAVGNNEENMDMTSPLELNSQYTNRKRPGLRERFSATLQRNLPGHSMLDRELTTDGQKNQESRFSPGMIMDRIMHLGVRTRLGKVRNSASKYGGQVTFKFAQTKGTFLDQIMKHKDTSGGVCESISAHWISAHAKGESIFDQLYVGGQKGKFHIDTLFSIKQLQMDGYLDDEQSTMTEYWLGTQGIQPNRQKNDNMNEHSSKIVGETGTRGTKDLLRAILDTGDKGSGYKKISFLGKMAGHTVAAYVDDQKGVTFFDPNFGEFNFPDKVSFSHWFTDDFWPKSWYSLEIGLGQEFEVFNYEPKEP
SEQ
ID NO:26(PAU_02097)
MVYEYAKTNDRKRKLSTQSDNYEEKSFSPVLDLSRNNQNTPNMEDEYETPQNFINRTGREKLFRAIRMVASNKRDPITKDQVSVPPDGNLFTELKDKHLDRAAEYKKLKTWPTHASIIATSPSANTPIAQHVSGDDALSPYISTGDKPGAVQNTVRNWNGIGPASERRLRPEKTWSPIIEIDVNKLPDTTKIFDLNKPNNTFFSTTNSDIAQNAFADKEVLISPEIPGLAITRVINDPEEIKQIANLNPSQSLIEKKNTIPEEKIIFEEKKSVPIHDSDADIPSSSFVFPKRKKPRNIRSRTDS
SEQ ID NO:27(PAU_02098)
MVFEHDKTVERKRKPSIQLGNDKEKSSEQALELPQSKQNNPLLHDLITSNNLRKEAAVFAKQIGPSYQGILDGLEHLHNLSGNEQLTAGFELHRRITRYLEEHPDSKRNAALRRTQTQLGDLMFTGTLQEVRHPLLEMAETRPAMASQIYQIARDEAKGNTPGLTDLMVRWVKEDPYLAAKSGYQGKIPNDLPFEPKFHVELGDQFGEFKTWLDTAQNQGLLTHTRLDEQNKQVHLGYSYNELLDMTGGVESVKMAVYFLKEAAKQAEPGSAKSQEAILLNRFANPAYLTQLEQGRLAQMEAIYHSSHNTDVAAWDQQFSPDALTQFNHQLDNSVDLNSQLSFLLKDRQGLLIGESHGSDLNGLRFVEEQMDALKAHGVTVIGLEHLRSDLAQPLIDKFLTSENEPMPAELAAMLKTKHLSVNLFEQARSKQMKIIALDNNSTTRPAEGEHSLMYRAGAANNVAVERLQQLPAEEKFVAIYGNAHLQSHEGIDHFLPGITHRLGLPALKVDENNRFTAQADNINQRKCYDDVVEVSRIQLTS
SEQ
ID
NO:28(PAU_02230)
MKGIEGVIMLSHDILPEKLLVSEKKHENVGSYFSDDIGEQSEQTEVSHFNLSLDDAFDIYADISIENQQELKNKDNNTNIWSSLGRGDDDHNLKKIINDAFKEKLPQLMEYRRKGYNVIGLDKEGIKKLEGMLKAVPPEIQQPTMKNLYSAAQELLNTLKQHPLLPENQDMIQQSNLVIRNLSDALEAINAVSKVNQVEWWEEVHKTNKAQSDRLIAATLEELFFKVKDKRLPGSNDDYCQQEREETERKIKDLLLYDGYQLTAEHFKFGRLRKSLLAESRVTRLKLAEYLEKKSVGILTAARDAKMYAMKILLAQTRNNGFNAKDLINAGQVNDRLLSFQQYARHIRAVDGEIDGIILSNPLVVACIKETNDEPAHIKIARAILPVSEELGTVSKVLRETKEKVQPSKPKEELNHPHQDWWNRGDELWKYIKKTSWNIKETSVHVTQMVGYEASKTASRAKHKLKESSYSESINGAVKGTALLLLDEIQQAENRIRQIPQFAWDVQEAVEQHSSVIQRTAYPDELPELSELLNEQLKHEEARWQAVKKQSRDKLQELIAPITRLAQEKWAQDLYFQLGEELRKERQDRWKDIQQFDEIMAEAVGQFAEMARELDSEAVRLAEHGHSGGKELQEKVAKWLRDLSKLKGKVKAGVAKITGTSLDNFSRSGMLARGMSEWAEDLKQSYLQETLQEGSAVAAELFERTLMEVVEENRTHFAKESDPEAERFLKRLALALKHAAENTTVYPPTPEEILAGSRSLPEDIRHWAEKKVVSGAISAAFRGGFKLVTGTFSLPVRVVIRGAKTGGTLYRGVRAINRSVRLGQGPATQVKSKFINQELSKTAFRLTLSLSPLVAWGMAASITAGRLYNEKDYPEKIIKNIVIDLPEELLWIGGYAGINAAIRAHAEKAIQQAIQHALDEQADKLALRINKEIAGKSADVNVEIIPQETSVSPAETAQSTPEPLSDFASTSQLTMPELIDIQDNNSAQQPKVRRKRDVSVESEISIDNLNIINANTREDKVNSEIKSELRSELKRFENSDANSPMSDVERAIFIDLFLYKNKYEVSESQQDYKNTWLKFRRELESQENKEIKEYLRFRSIIEAYEIYDKKRLDDDTIPEAGTIIKEVIDFFQKLKKENPITFMKLAEAMVKFQYYYEEEDENEDRYFKMAEIYYFLNKTENEKKSKTFHLDIIDKYPNENNRLLDEFFLNKNNNNPDLDEIIYKLQSMQEKYRESYEMLSKVENIHQVLSDDSKNEENIFLDNRIIAAQVFDGSINISLQDKKKWLNRYDQIRNEEGSDGWKLMHIESILINLRRINTAINLTAMKSESALLLIDKLLNFQKKARENILHISETPHEDFTSYSQFKTRKELGNDDSKYYAQFDNYKDNHDAEKEAKEILSQVVARASLSFSELFDKVESIKLFSFVYKNRDGGAPLAAPGRTVVIKFPGKDTGGLVISNLFLRNHVKRISTKEMEDLKPLTEGMYTRATQHRSLGSYYHIGSQSEHTNALEILSGMNKEELKTHLKKQGIWFGEPALFSNEYPKQENTGHLENTTLKNAIIGVSTIQNNAAANYLRSTMYESTGWEKLGDRFIPFYEIGRRKHYDREYEINSEQLTLDIITSIAIAYPAARGIVATIRSSAIPSILKSGLRGSALFKSLSLELGKMGFNASKVFGGAVYELIEPYPINSHLNRHNVFNKVKDTAWEFHTDVGLKGGGLKDFIDRFTKEPKEITISGYKFKRIKYNQENFDTMQRMALDYAYNPDSKGKIAQAQQAYKTGKEDYNAPQYDNFNGLSLDKKIERYISPDTDATTKGVLAGKMNESIKDINAFQTAKDAQSWKKSANKANKVVLTPQNLYLKGKPSECLPESVLMGWALQSSQDAKLSKMLMGIYSSNDITSNPLYKSLKELHANGNASKFNASATSISNINVSNLATSETKLFPTEISSVRVDAPKHTMLISKIKNRENKIKYVFYDPNYGMAYFDKHSDMAAFFQKKMQQYDFPDDSVSFHPLDYSNVSDIKISGRNLNEIIDGEIPLLYKQEGVQLEGITPRDGIYRVPPKNTLGVQETKHYIIVNNDIYQVEWDQTNNTWRVFDPSNTNRSRPTVPVKQDTNGEWFKHSETGLKGGGPIDDIRKYIARKSAIKIFNQSINYSATKWPPEPIDKNIHMIWIGTKNISEKNIKLSIDTAKKNPDYNTSIIYDSGISGHEGAKKFMLEKFQDSNVNIIDFRKKSYFSQLKQEPSFAYYEQVIAENKYAQASDILRLLVLKYEGGIYKDIDDIQVKGFGSLTFPKGIGVMREYAPEAGKATAFPNTPIAVTKNNPIINKTLDLAVSNYQRGEKNVLKLAGPDVFTQALYQEIPGLDSKVLNAQLYQLELAKRQALGVPLEKPKNFADEQLTSAEKEKINRPYQSIRGLSGYVENGADHSWAVDTNIPSTSTQTSTIVTPLAPKTEMLPPVPSSSTKSSTSAPVLQEKISYNLATDIDATDYLNQLKQKTNINNKISSPAGQCESLMKPVSDFMRENGFTDIRYRGMFIWNNATEQIPMNHFVVVGKKVGKDYVFDVSAHQFENKGMPDLNGPLILAAEDWAKKYRGATTRKLIYYSDFKNASTATNTYNALPRELVLESMEGKTFITSPNWYQTFKRTHNIHPEVTVSDPATFSLNYSVNPTAENLSPPPPPPIPSHGQVPKTVTPPPPPMRSPLSLSQPLERLPANKTKPIGFNPGENKASFSKLEEAGKHYYKDDKSRQAAPVNTMSDFDNRYLSHTTEAPAPSNVAHLAPGNIYNTKVTAKGAEKPAYDIYISKDGESLITSSSYKVDDITTDSKFGKPLPYSEIMFNSLKKSGVDPKNLKRSVQASIENKVTQDVISAIGTRIQRGQVIRVSPTENPDAFYTLLGTDNCKATLHMLNQHAEEFGHKVVTSIEFKGTGYLVMNIGTSTQTSTIVTPPPMPGTSQLVQ
SEQ
ID NO:29(PAU_02805)
MPNKKYSENTHQGKKPLIKSEANNEHAIDNSPLGIGLDLNSILGNNSASLSQIHDYSFWKENISEYYKWMVVVKAHLKQLDWTLKSMDSPESAGANIAKNIGTTTLQTLLNTGGSIAGGAIGGAIGSAIAPGVGTIAGMGIGALAGTGLNYLNDTAIEKLNEKLEIAYPYPKTRNMIFDINNYDKNPLIKAIKKKTKKDNLKVMAGSSLTSQLLGRITPIKIPAYKLADLAVSHHRALAGLSSDKARHILDFTNSIREVLNESHSDAVAFMRKNYGDNAMGLSGLSSKIKGDKLTLDTLARTRNKIENRINSINKQTLKLSSKNSNE
SEQ ID NO:30(PAU_02806)
MEREYSEKEKHKKHPIQLRDAIEQHAEETANNSLGLGLDLHQAINTPKVPKDNYNEENGDLFYGLAAQRGRYIKSVNPNFDPDKTNSSPMVIDVYNNHVSNTILNKYPLDKLGKLYGNPQKYAKDIKVTNSLQQDVAASKRGWYPLWNDYFKAGNENKKFNIADIYKETRNQYGSDYYHTWHEPTGAAPKLLWKRGSKLGIAMAASNEKTKIHFVLDGLNIQEVVNKQKGSTPLEQGRGESITASELRYAYRNRERLAGKIHFYENDQETIAPWEKSPELWQNYIPKNKSQNESSTPQRNNGALYRLGGPFRKLRASLRKRS
SEQ ID NO:31(PAU_02807)
MVHEYSINDRQKRHSFSSANPIDPEVTNRENSRHRFPKDNYNKGHGDLFYGLAPERGKYIKEANPKFDPNNPENAAMIIDVYNDEISRVILNNNANKISTNRLLNFIYNFRKNRLENLMKNPEKYAKDIKVKDNLRENISPKKIEKYPLWNDYFEAGIRNKKFNIAEIFKETASQYNSDYYHAWHIGGNSAPRLLWKRGSKLGIEIAASNQRTKIHFILDGLKIEDVVNKTKGPAPLKAGPGESITASELRYAYRNRARLAGRIHFYENGKETIAPWDKDPELWQKYTPKNRSGMEL
SEQ
ID NO:32(PAU_03332)
MLKYANPQTVATQRTKNTAKKPPSSTSFDGHLELSNGENQPYEGHKIRKIKGLRQHLADRSLNKGHISPLMNKGLLVGSKDVSIDIPVIAHRYDSSHQLTDAEPLKADSHSNHLDPFYGVIAGFRGDQVTSSESGSGSIGVHWGKNTLDSNIMGVNVVNGASGTVGIRIALKDIQHGSPVIVTSGALSGCTMVYSVKNGYFFAYHTGQKPGNNEWKTGRQGVVATYLSHQALSPDSEPMTVGEQNNDLVNIFANYDQSVITYMGKPGVLIDKMAENVGVFNYDEIKPEKPAIRAGYSYALLAKDDKGKVNVKVLSEDVIVSSGKQGNTVKAINSLKKRLL
SEQ ID NO:33(PAU_03337)
MPRYANYQINPKQNIKNSHGKSSSSDFSSGYLSFSNNSLDDPFIRQQVKREFIWEGHMKEIEEASRLGNFAVSFRAAGGPTLRALGKGAAAKGHDILEKTIKPGSINKAYPKDEASDVIKKVQEAGIEGYVGHWDKKTGRLLGIYMSSGHGLSDEQVNGKIYPIDLNNLEASLSALKAKENWAALPFTGDYDMHDMISFTGQPHSVPSNSSEERKIIDRINRLVARSDSNRPFGDIEHNVIRHGAQVSYPAFAMDKEKEEIKKHGGIVKAVAEPGEFPVAIVSKGKWTIANNIDELNQFYNSIGAKMKVSWKPGAENPGFVSNPQRPGMARFSRKR
SEQ ID NO:34(Plu1651)
MPNKKYSENTHQGKNPLMKSGANNEHDLQDSPLGIGLDLNSMLVNSSTSLSQIQDYSFWKENISEYYKWMVVVESHLKQLDWTLKSMDSPESAGTNVAKNMGVTALQSLLNTGSSIAGGAIGGAIGSAIAPGVGTIAGAGIGALAGTGLNYLNDTAMSKLSKKLEIAHPYPKTRNMILDINNYDKNPIIKAIKKNVNKDNLKVTAGSSLTSKLVGTVTSPIKFPAYKFAELAVSHHRALEGLSDDKARHILDFTNSIREVLKESHSDAVAFMRKNYGDNAMGLSGFSSKIKREKLTLNTLAKTKNEIENRINSINKQTLKVSSRSRNE
SEQ ID NO:35(Plu1671)
MLSTEKHNKDTKHPRNREKKFSIQPENSTQDDEDIKNNSLGVGLDLDQMIRNTSSTLTNAPQKPEDGYYYHISRGNNLQSFLQNGFKPQGSPGPTLSEEDFSRRKIGIIKLIYSIIATTINKNRKAKKISKDNFLMPQEFWHEFKNFYQNIPTQTNIDDQLLKKSITESIDKLDQNKFMEKHSDRKQTIINNEREAILQQDERINEIISSRAKMIQQREAENTEGYIYLAPHKNTLLEYMKHLQEEKNLFLILAVKEDIFTEKGLEQDPQEPHGAVRYKGALSTEELNFVNQEGQICAIPASIGEMDYGDFILNQQQVIDFCKK
SEQ ID NO:36(Plu1672)
MPINDLKKKFEISPQAAQAIGAPARSNSSKQAEHQTEHLELDTSKNRRDRKDLNAQATPNQQHTKKLETEVNNGGNKSKAQAHTPDLVMKKESSVTPNTRKSPNEKIKAEDIFHRYKDRFSPSDRELPFEIMNEITNNGIAFSSEKAPESHLDKVKDKKFTLRHYTSGNGQEKPTFNEIGSNFNLVNEGIKTLKRTQGSNTNEDDWNRLGNTAFTFFLLAIDGEVSDRKFLSNTTHFAEIDIENPAELKELGLDETEFFASPDLLHEKNLSQAPAVKGKLSDLKSLLLKQSGIKPVQLQSLGAKGILERIDSKFNGSLEIKIPGNVKVKEWKKVEK
SEQ ID NO:37(Plu1690)
MPNSKYSEKVNHSANGAEKCSIHSNQYNINNCTLGLGLDLNKKLRTGNERNIEGAQPFIPFPSKQKQYSTSPIAMADILNESALTSQPIITDLINPQKIKMSDGVKNILNNKEGGGDLVFKALQIKPSDETLPFNALKIVDTYQEEMPNKDMSISAYWAPQGGYVDIPAQPDISRHPQYVFTPNFSGCSFVVDKMNEDTLRVRHVQGGQEDVEYNNQNIDHGMGMITAMEFRDYGYHEADDKVIENTYGFAFLKFNQEKKQWQLHYQKIAAAPNIINIKTKSSWLPFSKPSIEADTFTFKNMKVPGYSRKNINNN
SEQ ID
NO:38(Plu1691)
MPKLTELLSRFENPIQNQPNHISKKNPISNSKVLNNSEEKTAPLELKHDDSKIKSQVSIPNLVKKNEKPAASNTPNNSHEKVKAEDIFNRFKSKFDPYDRELPFDIMNKITNNEIKFSSEKSKDDYLAKVKDKKFTLRHYTAGTGQEKPTFDEISSNFNLVNKGIKTLNRTQGSNTNEDDWNRLGNTAFTFYLLAIDGEVSNRKFLSNTTHFAEINIEDSEELKELGLDQAEFFASPDLLHEKNLSQAPAVKGKLSDLKSLLLKRSGISSVQLGRLDAKAILKSIDNEFGNSLEIKIPGNVKVNKWNKI
SEQ ID NO:39(Plu1712)
MPRYSNSQRTPTQSTKNTRRTSPSSNSSTEHLSLSNAPTNDSSVRQEVKEKFIWEGHWEGHMEAIEKASILGNFAVSFRAAGKPTLEALGKGAAAKGHDILEKTIKPGSIEKAYPENEASDVIKKVREAGIEGYVGHWNKETGRLEGIYMSSGHGLPNGQVNGKIYPIDLNNLEASLAPLKEKKNWAALPFTGDYDMHDMISFTTQPHSVPSNSSEEKKIIDRINEYIAKSDSNRPFEDIEHNVIRHGPQVSYPAFAMDKEKKEIKERGGIVKAVAEPGEFPVAIVSKGKWTIANNINELEQFYNSIGAKMKASWKPGAGNPGFVSNPQKPGMARFSRKK
SEQ
ID NO:40(Plu1713)
MFSTYSSKNDNQTINKINTEEKHENTETDNHLEINLEHTGKSKPDIEPKDVTTGTINAGTLLYKTTAIPEFLDNAKSLGLAEYEKRHKDIQDYLNLGKAEDAEKLKNKSQWAGQYFALEKSYDEYANEAPDSYNNLLKNAGKDLLENTEEVKVFLYTFKVTKDIKVLKPHNNSNSYYVGDTEGWEKAKEIMNDVQSQSEKNDNPFPELKNLEDKNFLLEELGEKGYAWMGPLHAKEGAEKGTEFSYELAISPNLLRQHLTLESEELLGTYKNRYGYWDKK
SEQ ID NO:41(Plu1714)
MKKTDEKYGQYEYKDEDITSYPIAWTNPDNGKIYIGINSPEYSHLNNKGESELNLAKIISTIIHESLHASSHQHKGLQSQTDTGADNLNYDEYVTDYFAREVYKQILPDKDYVANCFTKGLGGENKIWGGNIVEFMIQ
SEQ ID NO:42(Plu2400)
MVYEYDKTIERRRNPSIQLNNNEKSSEQALELSQNNPLLHDLITSNNLRKEAAVFAKRIGPSYQEILDELEHLHHLSGNEQLAAGFELHRRITHYLEEHPDSKRNTALRRTQTQFGDLMFTGTLQKIRHSLLEMAETRPEMASHIYQIAREEVKGNTPGLTDLMVRWVKEDPYLAAKTGYQGKIPNDLPFEPKFHVELGAQFDDFKKWLDTAQSKELLTHTRLDEQNKQVHLGYSYNELLDMTGVESVQMAVYFLKEAAKQAEPGSTKSQEDILLHRFANPTYLAQLEHSRLAQIEAIYHSSHDTDVTAWDQQFASDALTQFNHQLNNTVDLNSQLSLLLKDRQGLLIGESHGSDLNGLRFVEEQMEVLKAHGVTVIGLEHLRSDLAQPLIDKFLASGNEPMPAELAALLKTKHLSANLFEQARSKQMKIIALDNNSTTRPTVEGTQHGLMYRAGAANNVAVERLRQLPAGEKFVAIYGNAHLQSHEGIDHFLPGITHRLGLPALKVDENNRFTAQVDNINQRKRYDDVVELPRIQLTS
SEQ
ID NO:43(Plu2401)
MEHEYSEKEKPQKCPIQLRDSIEHDKEDINTTTPLELNSQYTNRKRAGLRERFSTTLQRNLPGHSMLDRELTTDGMKNQESRFSPAMIMDRMMHFGVRTRLGKVRNSASKHGGQVTFKFAQTKGTFLDQIMKHKDTSGGVCESISAHWISAHAKGESIFDQLYVGGQKGKFHIDSLVSIKQLQMDSYLDDEQSTMTEYWLGTQGIQPIMQKNDVDEHSSKVVGQTGNKGTTDLLRAILDTGDKGSGYKKISFLGKMAGHTVAAYVDDQKGVIFFDPNFGEFSFPSITSFSRWFTDDFWPKSWYNLEIGLGQQFEVFNYELKKS
SEQ ID NO:44(Plu2514)
MYDSKKKNSEPTTKKKFERSNYSQWDDSINHYEDMNRARIKNRNDILTTVDYFGEKKKTMHTFEYQSDIKHDTNFNNKNKSLFESFAASFVLQNPSFFSGVIDKLSKKLFNIISKIDERNNFQKKLYDFIEKDTSPEGQFGRFTLGKNEILNVLQVKSDTPQLFVKKMLLIKSLGAFIIDFSSKDIGNYDFIFDGKGREVNDIIEKNRPTNLFKVRGRTNIKSSQHRSDIGILDTPTFDSLTEEQKSFLTIPELTKRRPLFRTFTHELDAEDKRVVESVFVNRTFDCDSPLIGSVSGSTSCVLVAADILFPDMTMVERKKLAIATFAFLVGGGYHSATEVFDVAYPGLDLNKEIEELIENNPIQENAGVATLRQLIGNSGF
SEQ ID NO:45(Plu2515)
MPISNLAKESEVRAVKDIPCKNIETDNHLEIGLSSGLSRSKDTSKFKKNSINTIKLIDDIIALHNDPKGNKLLWNDNWQDKIINRDLANIFEKIDESVSELGGLEMYQEMVGVNPYDPTEPVCGLSAQNIFKLMTEGEHAVDPVEMAQTGKIDGNEFAESVDQLSSAKNYVALVNDRRLGHMFLIDIPSNDQETVGYIYQSDLGQGALPPLKIADWLNSRGKDAVSLNKLKKLLSREFNLLSDDEKRALISETLDIHKDVSNVELDRIKRDRGVDIYLTEYDVNNFYENIETLKSKLSNYDKKLSKPK
SEQ ID NO:46(Plu1649)
MLANVLPNLASFLKYEKETPLFFIEDGFNFQNLNPGRVPLIKTPEQRKAGDTQSPAFLCSGVILRGTIHSNDYKFWQPSPSSIKSGGVSFSYLRKDAKFKRLAYGYKNGFIIFPEHIAPEDRVDFSVLCAFPIDGYTNERANQGCGENITKAKDKGKSCQEQNVTNSDDWIKNYRKVNSQDFFQCGFNVTKDVNNPAIAFYQMLESIKKLPRTPNTPPKQNEIRISTWEESDPNKLPIEALFYSENSGLADAQKDQRDYKNATGKFLPIVKMLLPRTLNEDALFKFNIKDQVINP
前导序列(例如，SEQ ID NO:47-92分别对应于SEQ ID NO:1-SEQ ID NO:46的氨基
酸1-50)
SEQ ID NO:93(不对称光杆状菌菌株ATCC43949PVCPnf操纵子，pvc1-pvc16；例如，
对应于GenBank登录号FM162591.1的序列的基因PAU03353至PAU03338)
TGAGAAAATTCCATTTAATAGTGTTTCAGGATTAGATATTAGTTATGACACCATTGAATACCGAGATGGTGTTGGTAATTGGTTCAAAATGCCGGGTCAGAGTCAGAGCACTAATATCACCTTGCGTAAAGGCGTTTTCCCGGGGAAAACAGAACTGTTTGATTGGATTAACTCTATTCAGCTTAATCAGGTAGAGAAAAAGGATATTACCATCAGTTTAACTAATGATGCAGGTACCGAATTATTAATGACCTGGAATGTTTCTAATGCTTTTCCCACTTCATTGACTTCACCTTCATTTGATGCCACCAGTAATGATATTGCAGTACAGGAAATTACGCTGATGGCAGATCGGGTGATTATGCAGGCTGTTTGAAGCATTGATATTTAATCATCTCATATAAGGGAACTTTTATGACAACCGTTACCAGTTATCCTGGCGTTTATATTGAAGAATTAAATAGCCTGGCCTTGTCAGTTTCAAATAGCGCCACAGCGGTTCCTGTTTTTGCTGTGGACGAACAAAACCAATATATTAGTGAAGATAATGCAATCCGTATTAATTCGTGGATGGATTATCTTAATCTGATTGGCAATTTTAATAATGAAGACAAATTAGATGTTTCTGTGCGTGCTTATTTTGCCAATGGAGGTGGATATTGTTATCTCGTCAAAACAACGAGTTTAGAAAAAATTATTCCAACCTTGGATGATGTAACCTTATTGGTTGCTGCGGGCGAAGATATTAAAACGACAGTAGATGTTTTATGTCAGCCAGGAAAAGGGTTATTCGCAGTCTTTGATGGCCCTGAAACAGAGTTGACTATCAACGGTGCGGAAGAGGCAAAACAAGCCTATACCGCCACACCATTCGCTGCGGTTTATTATCCTTGGTTGAAAGCGGATTGGGCTAACATAGATATTCCACCCAGTGCAGTGATGGCGGGAGTTTATGCATCGGTGGATTTATCCCGTGGTGTATGGAAAGCGCCTGCCAATGTTGCGTTGAAAGGGGGCCTGGAACCTAAATTTTTAGTCACGGATGAATTGCAGGGTGAATATAACACTGGCCGCGCTATCAATATGATTCGTAATTTCAGTAACACAGGTACTACGGTTTGGGGTGCAAGAACCCTGGAAGATAAAGACAATTGGCGTTATGTTCCAGTGCGACGCTTGTTTAATTCTGTGGAGCGGGATATCAAGCGTGCCATGAGCTTTGCTATGTTCGAGCCTAATAATCAGCCTACTTGGGAGCGGGTACGGGCGGCGATTAGCAACTACCTTTATAGCCTGTGGCAACAGGGGGGATTAGCTGGCAGCAAAGAAGAAGACGCTTATTTTGTGCAAATTGGTAAAGGTATAACGATGACACAGGAGCAGATTGATGCAGGGCAAATGATTGTTAAAGTCGGTTTGGCTGCTGTACGGCCTGCGGAATTTATCATTCTCCAGTTTACGCAAGATGTAGAACAGCGTTAATCATATGATTATGAGGAGTTATCATGTCTGCTATTCTGAAAGCGCCTGGCGTTTATATTGAAGAAGACGCTTCCCTAGCGTTGTCTGTCAGTAACAGCGCGACTGCCGTGCCTGTTTTTATCGGAAAATTTACTCCGACAGTGGTTGATTCAATCCAAGTCTGTACCCGTATCAGCAACTGGCTTGAATTCACTTCCTCTTTTTCCCTAGCTCCAACAGTTGAGATTGTTGTCCAATCTAACACTGAATCTGAATCTGAATCTGAAACTTACCACTATATTGAGACAATCAATTTATCTCCAGCTGTGGAAGCATTGCGACTCTATTTTCAAAATGGCGGAGGAGCTTGCTATATCTACCCATTAAATGATGCTGAAGATGAATTGGTTCTGGCGGCCATACCAGAAGTCATTGAACAGAAAGGTGATATTACTCTGTTGGTTTGCCCGGAACTCGATCTGGATTACAAAACTAAGATCTATGGCGCAGTGAGCTCACTGTTGAATGATAACAAAGTGGGCTATTTCCTGATTGCGGATAGCAATGATGGAGAATCTGTGTCAGGAGTATGGAATAGTGCTAAGGCCGCCGCCTATTATCCCCAGTTGGAAACTAACCTAAAATTTTCCACGTTGCCTGGGGATAAGGACATTCGTATCAGCGGTTATCAGGATGATGATGAAACACATAAACCGAAAAACTTGGATGAGCTCAGGACAATCAACGAGGCGTTGGCACAGGATATTGATGCAAGATTGCTCGAGGAGAAACAACGTGCTGTCATCATTCCGCCAAGTGCTGCCATTGCGGGCATTTATTGCCAAACGGATAATCGTCGCGGTGTTTGGAAAGCGCCAGCCAACGTTGCGCTCACAGGGATCGGGAGTTTGCTTGATAAGGTAGACGATGAACGGCAGGGAGAGATGAATGACAAGGGAATCAATGTCATCCGTTCATTTACCGACCGTGGTTTTATGGTCTGGGGAGCCCGTACTTGTGTGGACGCTGCCAACATCAGCTGGCGTTATATTCCTGTTCGTCGCCTGTTCAATTCCGTTGAACGAGATATCCGCCAGGCGCTGCGCGCTGTGTTGTTTGAAACTAATAGTCAGCCTACCTGGGTACGTGCTAAGGCTGCCGTTGATCAATATCTTTATACCCTTTGGCAGAAAAATGCATTGATGGGTGCTCGCCCGGAAGAAGCTTATTTTGTGCAAATTGGTCAGGATATCACCATGTCCGAGGCTGATATTAAACAGGGTAAGATGATCATGACTGTTGGTTTGGCAGCAGTGCGGCCAGCTGAGTTCATCATTCTGCAATTTACGCAGGATGTTGTTCAGTAATCTCCATGACTAAACGCCAGGCACTGTATTGACAGTGCCTACTCTAACCATCTTGGAGGAGGTGATGATGATGGAGAGACTCCAACCGGGTGTGACTTTAACAGAAAGTATAATCACGATGGGTCAGCAAGAGATACCCAGTGCTGTGCCGGTGTTTATTGGTTACACCGTTCGTTATCCGGAACAATCGGAAGCATCAGTCCGTATCGACAGTTTGGCCGAGTATACCAGCCTGTTTGGTGACGACCATGTGATGATGTTTGCTGTCAGGCACTATTTTGATAATGGCGGGCAACAGGCATTTGTTTTACCCCTGAAGGACAATATGCCATCAGTGGAGATGACCACAGCTGAAGCGGAAAATCTGATAGCCGCATTGCGCTCTGCTACGGTTAGCGAAGCCATTGGTGGGCATAGTCAGATTACACTGATTTTGGTACCGGATATGGCTCGGCTTAATGACAGTGATATTGATGACTCCTCAACCCAGGTAAGCCTGTGGTCCCAAGGCTGGGAGGCGCTGCTGCAATTGAGTCAGGTTAGGCCCAACCTCTTTGTGCTGTTAGATGCGCCGGATAATGTTGAACAGGCGCAGAAGTGTATGACAACGCTATCGTCAGATTATCGTCAATGGGGGGCAGCATATTGGCCTCGTCTGGAAACTACCTATCAGAAAGAAATATCTGGCAAGGACAATGAATCTCAGGGAATTTTCCAGGGGACTGTTCTGTCACCCACAGCCGCGGTCGCAGCGGTAATTCAACGCACGGATAACGACGCGGGTGTTTGGAAAGCACCGGCCAATATTGCCTTATCCCAGGTTATTCGACCTGTTAAATCTTATCTTCAGGGAAGTGTACTGTTTAACAGCAGCGGCACTTCGCTCAATGTGATCCGCAGTTTCCCAGGTAAGGGCATACGGGTATGGGGATGCCGCACTCTGGAAAACACGGATAATACGCAGTGGCGCTATCTGCAAACACGTCGGCTGGTTTCCTATGTAACAGCGCATTTGACCCAATTGGCTCGCATGTATGTCTTTGAGCCAAATAATGAACTTACCTGGATGAAGTTAAAAGGACAAAGTTACAACTGGTTACGGCAATTATGGTTGCAGGGTGGCTTGTATGGTTCACAGGAGGATGAGGCATTTAACATTCTGTTAGGCGTAAACGAGACGATGACTGAGGATGATGTTCGTGCAGGAAAAATGATCATGAAAGTTGAGTTGGCTGTGTTGTTTCCTGCCGAATTTATTGAGATCAGTTTGGTGTTTAATACCCAAACAGAGGCGCTGTCTTAAGAAGGAAAAAGTACGATGAACGATTATTACACACCCGTGGTATCCCATCGTTTTATGGCGAGTTTTATTTTTAACCGCATTCCCGATCCGCTGGATATTCGTTTTCAGCGTATCTCTGGCCTTAGTCGGGAACTACAGGTGACTCAGTACAGTGAGGGAGGAGAAAATGCCCGTAATAACTATTTAGCTGAGAAAATCCAACACGGTACGTTGACTTTGGAACGGGGCGTGATGACAGTCTCGCCATTGACCTGGATGTTTGATCGGGTATTGAGTGGTGAAAAAATCGCTTATGCCGATGTGGTGGTGATGCTACTGAATGAAAATTCACTGCCATTGTCCAGTTGGACGTTGAGCAATGCGCTGCCGGTACGCTGGCAAACCAGCGACTTTGACGCTAACAGCAATGCCATATTGGTGAATACCCTTGAATTGCGTTACCAGGATATGCGCTGGCTTGGAGTCAAAATATGACAGTAGAAATCAGAGAGTTACTTATCCAGGCAAAGGTAGTGCCATCAACACGACCGACTGAATCAGAACGGCAAAACCATTCTTTGATACAGGAAAGTCTGGATGAGGCGACTTGGGTGGAAACGATAAAACGCGAAGTGTTGGCCGCATTACGCGATGAGGAAGGGTGGCGTCCATGAGTCTGATTGAACGTGGTTTAGCTAAGCTGACAATTAATGCTTATAAGGATAGGGAAGGGAAGATACGGGCAGGAACGTTGCAGGCCATGTATAACCCTGACTCCTTGCAACTGGATTACCAAACGGATTATCAGCAATCCCAAGCGATTAATAGCGAAAAGCAAAGTAGCATTTATGTACAGGCCAAGCCCGCAGGGTTATCACTTGAATTAATTTTTGATGCCACGATGCCGGGTAACAAAACCCCCATTGAAGAGCAGCTCATGCAGCTCAAGCAACTGTGCAGTGTGGATGCAACCAGTAACGAGACGCGATTCCTGCAAGTTAAATGGGGCAAAATGCGTTGGGAAAGTCGGGGTTACTTTGCTGGCAGGGCCAAGAGTTTGTCTGTGAATTACACTTTGTTTGATCGTGATGCGACTCCCTTGAGGGTACGGGTAATATTGGCATTAGTGGCTGATGAAAGTCTGGTGTTGCAGGAGACTGAACAAAATCTGCAATCTCCGGCAAAAATCGCATTACGCATACAGGATGGGGTATCTCTGGCTCTGATGGCAGCCAGTACGGCATCAACATTGTCAGGCGGTGTGGATTATCTGACGCTGGCCTGGCAAAACGGTCTGGATAATCTCAATGGGTTCGTTCCGGGTGAAATATTGCAGGCCACCAGGGGAGACGAATCATGAGCCACCAACTGAAAATTATTGCAGATGGTAAGGCACTGTCACTTTTGGCCGCGGTAGATGTGGACACCTGTTATCGGGTTAACAGTATACCTTCTGCGACATTGAAACTGAGCGTACCGGATAGGCCACTCTCTTCTTTCAGTCAGACGGATGTTCAGACAGAACTGGCCCACTGTCAGGTAGGGAAAACCCTGCGTCTGGAATTGATTGATGGTAGCAAAAAATGGGTGCTGTTTAATGGTCTTATTACCCGTAAGGCTCTGAGAATTAAGAATAAGCAATTATTGCTCACTCTGGTTGTCAAGCATCGGTTGCAACTGATGGTGGATACCCAGCATTCACAGCTGTTTAAAGACAAAAGCGAAAAAGCGATCTTAAGCACGCTATTGAATCAGACCGGAATCAATGCTCGCTTCGGAAAGATAGCGGCGTTAGATCAAAAGCATGAACAGATGGTGCAATTTCGTTGTTCAGACTGGCATTTTCTGTTGTGCCGACTGTCGGCAACCGGTGCATGGTTGTTACCTGCCATAGAAGACGTTCAGTTTGTTCAACCTGATGCTCTGAAATCAAACTCAGCCTATACCTTGAAGAGCAGGGGGGATGAGAACAAAGACATCGTTGTCAAGGATGCTTACTGGCAGTTTGACAATCAAATCAACCCCGCTTTGCTGGAAGTCAGTGGCTGGGATATCAGTAAGCAGCAGGTACAATCAGGCGGTCGCTACGGAAAAATCGCGTTGGGTAAGGCGGCACTCTCTCCTGATGGATTGGCATCCCTTAATAAAACGGGTTGGGACATTTGTTATAGCAGTCCGTTAACAACCCAGGAAAGCGGTTATCTGGCACAGGGATTATTGCTTAACCAGCGCATTTCTGGGGTGACAGGAGAATTTTTGCTCAAAGGAGATGGGCGTTACCAGTTGGGAGACAACATTCAGCTGACTGGATTTGGTTCACAGTTAGATGGTACGGCAAGCATTACTGAGGTTCGCCACCGTCTTAATCGGCGAATTGATTGGGAAACCACGGTGAGCATTGGTTTACAACATGAATATTTGCCGATATTACCTGATGCTCCCGAACTACATATTGCGACAGTAGCGAAATATCAGCAGGACAGTGCGGTGTTAAACCGTATCCCCATTATTCTGCCGGTACTGAATCGTCCCAATGAATTTTTGTGGGCCAGATTGGGGAAACCTTATGCTAGCCATGAAAGCGGTTTCTGTTTTTACCCAGAGCCAGGTGACGAAGTTATTATTGGTTTTTTTGAAAATGATCCGCGTTATCCAGTTATTTTAGGTGCTATGCATAATCCGAAAAATAAGGCCCCTTTTGAACCAACCCAAGATAATAGGGAAAAAGTATTGATCGTTAAAAAAGGTGAAGCGCAACAACAATTAGTCATTGATGGCAAAGAGAAAATGATCCGAATTAATGCGGGTGAAAATCAAATAATGCTTCAGCAAGATAAAGACATTTCTCTGTCAACGAAAAAAGAATTAACACTGAAAGCGCAGACAATGAATGCCACGATGGATAAATCATTGGCAATGTCCGGGAAAAACAGTGTTGAAATCAAAGGCGCAAAAATTAATCTTACCCAATGAAAGGTGACGATGAATGGAAAATCAAATACTGACACAACTCTATGGTCGTGGTTGGGCTTTTCCTCCGGTCTTTTCCCTTGAAAAGGGGGTAGAGATGGCTGAAGGGGCGGAAGATGTGAGACAAAGTTTGCAGATTCTGTTTAGTACTGAGCCGGGGGAACGTCTTATGCGTGAAAATTATGGCTGCGGATTAAATGATTTTATGTTTGAAAATATCCGCAATGAACTTATTGCTGAAATTGAATCCCATATCCATGACAACGTATTACGATATGAACCCCGGGCTGATATGACTGATATTCAGGTTCGTCAATCCCCTGGCATGGGGAATACTTTGCAAGTGCAGGTCATGTATCGCCTGAGAGGGAGTGATATCAATCAACAAATCCAGGGAGTACTTGCACTGAGTGAAGGCCGGGTGACGGAGGTAGTATGAGTGAAGCGATTGTGGTGGATGGTGACGTGTTACAGTTTGATCCCAACTTTGGCAATCGGCAGGTGACGGTTCCCAGCCCAGGAAAAATTAGCGGCACAGGACATGCGCAGGTAAGTGGAAAAAAAGTGTGTATTCTGGGGGATGAGAAACAGGTCAGGGTTTCTGCAACCTATATTACAACAACACATACTACGCCGGGAACAGGAACCATTACTATCAGTGCTCTGGATGCTGGCCAGCAGGCCCTTCAGTGTACCAGTGGGGCGGCTTTAATTATCAAGGGGCAGCAATTTACGGCGATGTTTACGCCTGAATTGCCAGCCATGAATAATACAGTGACTCCGCCACAACCGGATGTTACGACACCTTCATCAGGAAAAGGACGTTTTATCACTCAACAAAATTTTGCTACCGTAAATTAGAGTATTGACTGAATTAAATAGAATTAACGAAGGTGTAAATAATTATTTATTTGCTGACGAATCGCTGTGACAAATAAACACAGGTGATGTTATGGAATTAAATGAGTTAACTAACAAATTGTCAAATTTGGTGCCAATGACCGATTTTAAATTAGATAATCGAGCCAGTTTGCAATTGCTTAAATATATTGAAGCGTATACGAAGATAATACCCTTTAATTCTGGCGATAAATATTGGAATGACTTTTTCTTTATGTCAGGAAATACGCCAGAGAAACTTGCAAAATTATATCAGAAAGAAATAGAACCCAATGGGGAGTTATTACCTCAGCAGGCTTTTTTGTTGGCGGTTTTGCGTTTATTGGAAACACCAATATCCTTATTAAATGTATTACCTGCTGCTCATCGTGAGCTCTATTATCGGGAGCTTTTAGGCTTGTCTTCCCATGCGGCACAGCCTGATCAGGTTGCTTTATCTATGGAACTGAATTCGACAGTGATGGAACAGCTGCTCCCTGAAGGAACCCTGTTTGAGGCTGGTCAGGATGAACAAGGCAATGCATTGCAATATGCCCTGGATGCCAGTTTGCTGGCTAATCGTGGATATATCAGTGACTTGCGCTGGTTACGGAATGACGGGGAAAAGCAATGGGTTACTTCTGCTCCATGGGATTTACAGGCACAGGTGTCACTGCCGTCTGATGGGATACGATTATTTGGTAAGACAAATAGTGATCAGCAGGTATTTGGTGGGGTGTTGATAACGTCATCACTTCTGGCGATGGAAGCGGGGATAAGGAAGATCATTGTTACTTTTGAGCAGGAGATGAACACCCAAGAACTGGTGGCACAGGTCAGCAGTGGAAATCAATGGCTAACATTGACGTCTGAGGTAAATAAGAAAGAGGTCACACTGACACTGTCAGACAAAGAACCGGCAATCAGTGCGCCAGAGGATCTGGATAATCTCTTTTTCACGCAACCGGTACTCAGGCTACAGGGAAAGGATAGTCAGGCACTGCCGGAGGTGACGGGTATCAGCGTTTCGGAAAAGGATGATACTAAGGATACCTCTTTTGAGATGTATCACTTAACACCATTTGGTTATAGCAGTGATATAGAGCCATTGGAGGAAAATCCAGCGTTATATTTAGGCTTTACTGATGTAAAGCCAGGGCAAACACTGGCGCTGTATTGGAAATTAAAATCCCCGCAGCAACCAACCGTTTCCTGGTATTACCTGGATCAACATAATCAATGGGCTGAATTGGATTCATGGGTCAGTGATGGAACCCAGAATCTGTATCAGGATGGTACTTGGCACGTTGAGTTGCCTGTGGATGCATCCAATCAGGCAGAGCAGATGCCAGTTGGACGCTATTGGTTGCGGGCAGTGGTGGAGGTACCCGCTCATGAGGGGGCGTTGGGGAAGGCTCCTTGGCTATATGGTCTAATCTATAACGCCATGACGGCAACCTTGGTTAATGTAGATAGCATCAGTGACAGCCATTTCTTAACCCCTTTGCCTGCCAGCAGCATACAGCGGCCCGTTGAACCCATCATTGTGTTGGCATCGGTCAACCAGCCTTGGGCATCATGGGGTGGACGTATACCTGAATCCTACAGTGCCTTTTTTGAACGGATAGCTCAAAACCTGTCTCATCGAAACCGGTCCTTAACCTGGGGAAATATGGTGACATTACTCAAAGAGCGTTATGTCAGCATCTTTGATGTTAAGTATCCAGGTAATGATGAACTCACCAGAGTGCCAGCATTGGAGCAGCAGCAACTAACAGTGATTCCAGCAAACCGGTACAACGATAGCGATGATTCTCTGCGTCCGGTACTGAATCCTGCTCGTCTGCAAGAGATGGCTGATTGGTTGCAGCAGAAAGACTCTCCCTGGGCCTCTATTGAGGTCAGGAATCCAGAATACTTGGATGTGAAAATCCATTACGAGGTGATTTTTAAACCTGATGTGAACGAAGATTTTGGCTATCGCCAGCTACAGCAGCAACTGTGTGAGGTGTATATGCCTTGGAGCATAGATGAGCAGCGGCCCGTTGTATTGAATAACAGCATTAATTATTTCCAGTTGTTAGCCACTATTCAACAGCAACCGCTGGTTGAGCGAGTCACTCGTCTGACACTACATCGGGCTGATTCTTCTGATGAGAGTGATGGTACAGCATCTGTGGAAGCCAAAGATAATGAAGTGCTTATTTTAGTCTGGGAAGAGGACGATAATCTGCAATACCGAGGAAATGACTATGAGTAATCAGGATGCACTGTTTCATAGCGTTAAAGACGATATTCACTTTGATACCTTGCTGGAACAAGCTCATCAGGTGATTGAAAAACAGGCTGAAAAACTGTGGAGTGATACGGCAGAGCATGATCCGGGTATCACATTTTTGCAGGGAATCAGTTACGGTGTGTCAGATTTGGCTTACCGACATACATTACCCCTGAAAGATTTACTGACTCCGGCGCCGGATGAGCAGCAGCAAGAGGGAATTTTTCCTGCCGAATTTGGCCCGCATAATACACTGACTTGTGGGCCGGTGACAGCGGATGATTATCGCAAGGCATTGTTAGATCTACACAGCAGCGACAGCCTGGATGGTACTCAGCAGGATGAGGGGGATTTTCTGTTCCGGAGTGTGCAACTGGTGCGTGAACCGGAAAAACAGCGTTATACCTATTGGTATGATGCAACCAAGAGGGAATATAGCTTTGTCAACAGTGAAGGGGCTAAAGAGTTTACCTTGCGGGGGAATTACTGGTTGTATCTGGAACCAACCCGTTGGACTCAGGGTAATATTGCCGCTGCTACCAGACAACTGACAGAATTTTTGACTAAAAATCGCAATATTGGTGAATCTGTCAGCAACATTATCTGGCTACAACCGGTTGATCTGCCACTGTTGCTGGATGTTGAACTGGATGATGATGTAGGTGCACAGGATGTCCCCGGTATTTTTGCGGCGGTGTATAGCACCGCAGAGCAGTATCTGATGCCTGGAGCACAGCGTTACCGTACGGAAGTACTGCAAAATGCTGGGATGAGCAATGATCAAATCTTCGAAGGTCCATTATTGGAACATGGCTGGATACCAGAGCTGCCGGCAGCCCGTGATTATACTCAAAGGCTCACTCTCAATCTTAGCCGGTTGGTAAATAGTCTGCTTGAGATTGAGGGCATTAAACATGTGAATCGTCTTCGTCTGGATGATAGCTTCGATAAAACTGCTATTGAACCCGTTAAGGGGGATACCTGGTCGTGGTCGATCAAAGAGGGCTATTATCCACGTCTTTGGGGAGAAGACCCACTTAACCAATTGGCGCAACAAAATGGCCCGCTTAGGGTGATAGCCAAAGGAGGGATTAGCGTCAGTGTGAGTAAAGAGCAAATCCAGGCCAGTTTACCCAGTCAATCACTGATTCAAAATGAGCCGGTAATATTGGCTTACGGCCAGCACCGTGACGTTGGCAGCTATTATCCCGTCAGTGATACTTTGCCGCCTTGCTATGGACTACAACATTCTTTGTCTGAAAGTGAACACTTATTGCCACTTCATCAATTTATGTTGCCATTTGAACAATTATTGGCCTGTGGTTGTCAACAGATAGCCATGCTCCCGCGGTTACTGGCTTTTCAGCGCGAAGGTTATGAGGTTTGGGGTGATCAGTGGCCCTTTAAGTCAGGCTCAGTGAATGATGACGCCCATCAAGATTATGCCCCTGCATTAAAGGATTTGTTAGGACAGATTGCGCTGGATAGTGATCATGAATTGGATATTATTAATTACTTGCTGGGTTACTTTGGCACACAGCGGGCACCGCGTACCTTTACGACACAACTCGATGATTTTCGTGCGGTCCAACAGGGTTATCTGGCCCAGCAACCGACATTGACTTACCACCGCTCCAATATTCGTATCGATCAGGTATCGTCGCTACAAAAACGTATTGCTGCTCGCATGGGGCTGGGCGGTGAGTTGTTTAAACCTCAACCGGATCTGAGCCAACTGCCTTTTTATTTGATTGAACATCGAGCGTTGCTGCCAGTCAAACCCAATAGTCAGTTTGATAAGGAACAGAAACCAGCCTCGGTGACAGAGGAGGGGGGCAGCCAAACAGGTCAACATTATGTGGTCATTGAACAGAAGGGCATTGATGGCAAGCTGACACAGGGGCAAGTGATCAATTTAATTCTGTATGAAGGAGAGCAGGGAGAAACCCAATTTACGATACGCGGTCAGATGGTATTCAAAACCGAGGGGGATAAGTTTTGGTTGGATGTGAATAATAGTGCGCAACTGGAATATAATCTGGCGCGGGTAATGACAGCAGCCAAGGCGAGTAAACTCTTTTGGCAAAACAGCCCGGTATGGATGGAGGATATGGGCTATCGTCTGGCCTATGCTAGTGACCAATCCTCATTGCCTGTGAATCAACGGCGCTTGACCCGCACAGTGCAAACTCCATTCCCGCCGATGGTTGTTGTAGGTAGCGAAATCACCCTGTTAAAGCAGGTGGGGATAGTCAATTTAAAAAAAGCGGAGTCAGAAAAACTTTATGCAAAAGTTGTTAGCTTTGATCGCATTGAAGGGACCTTGATTATTGAGCGTTTGGGTAATTCCACTCTGGCTTTTCCTACCTCGGAAGAGGCGTGGCGGTATAGTTGGTATTTTTCGGGGGAGAAATATGAAAGGACTGACCGCTTTTCATTTGTGATTAGCGTAGTAGTGAACAGTGACTTAATTAAATTGCCCGGTGTTGATCCCTATAAATTGGAAGAATGGGTGAAAGAAACGATTCTTACCGAATTTCCAGCTCATATTTCTATGATTATCCATTGGATGGATCGGGAAGCCTTTTTAAATTTCGCCAATACCTATCAGCGTTGGCAAAATAATGGTACGCCACTGGGGGATGCGGCTTATTCCATTCTAGAAAGTTTGACACTTGGTAAATTGCCATCTGCCTTAAAAGGTGTTGGCACAATGCGTATTGCCACATCTAGTCAAAGAGAAGAAGTGGTGGGTAGTAATGGTGATCAATGGAATACAGATGGAATAACCCAGAATGAATTATTCTATGTTCCTAAAGAGAGCTAGGAAAAATAAATATCTGCCACTAATGATGTTGAATTAAATATGTTTTCTGGAGTTAATCATGAACGAAACTCGTTATAATGCAACTGTACAAGAACAACAAACATTATCTAATCCAAAAGCTGTTGGACCTGACATCGATAAATTAAAGGATAAATTTAAAGAGGGCAGTATTCCCCTGCAAACCGATTTCAATGAGTTAATTGATATTGCCGATATTGGACGTAAAGCCTGTGGTCAAGCGCCACAACAAAATGGCCCAGGAGAAGGATTGAAATTGGCTGATGACGGTACGCTTAATTTAAAAATAGGCACTTTTTCCAATAAAGACTTTTCTCCATTAATATTAAAAGATGATGTTTTATCTGTAGATCTTGGTAGTGGTCTGACTAATGAAACCAATGGAATCTGTGTCGGTCAGGGCGATGGTATTACAGTTAACACTAGCAATGTAGCTGTAAAACAAGGTAACGGAATTAGCGTTACTAGTAGTGGTGGTGTTGCCGTTAAAGTTAGTGCTAATAAGGGACTTAGCGTTGATAGTAGTGGTGTTGCAGTTAAAGTTAATACTGATAAGGGAATTAGCGTTGATGGTAATGGTGTTGCAGTTAAAGTTAATACTAGTAAAGGAATTAGCGTTGATAATACAGGTGTTGCAGTTATAGCTAATGCTAGTAAGGGAATTAGCGTTGATGGTAGTGGTGTTGCAGTTATAGCTAATACTAGTAAAGGAATTAGCGTTGATGGTAGTGGTGTTGCAGTTATAGCTAATACTAGTAAAGGAATTAGCGTTGATAATACAGGTGTTGCAGTTATAGCTAATGCTAGTAAGGGAATTAGCGTTGATGGTAGTGGTGTTGCAGTTATAGCTAATACTAGTAAAGGAATTAGCGTTGATGGTAGTGGTGTTGCAGTTATAGCTAATACTAGTAAAGGAATTAGCGTTGATAGTAGTGGTGTTGCAGTTAAAGTTAAAGCTAATGGCGGAATTAAAGTAGATGCTAATGGTGTTGCAATTGATCCTAATAATGTACTCCCCAAGGGAGTGATTGTAATGTTCTCTGGCAGTACTGCACCAACTGGTTGGGCGTTATGTGATGGCAATAATGGTACACCAAATTTAATCGATCGATTTATTTTAGGTGGGAAAGGGACTGATATTAATGGAGTGAGTACTAATACAGCTTCAGGTACTAAAAATAGTAAGTTATTCGATTTCAGTTCTGATGAAGCTACATTAACTATTGATGGTAAAACACTGGGGAGAGCATTATCGTTACAGCAAATACCTAATCATGCACACTTTAGTGGAATAATTATGGATACAGAGAAAGTTAATTATTATGGAAGTAAAAAAATCACAACAAATGTGTGGGGTGTAACAACAGGAGATAATACTTCAGTACGATATATTTATAAGTCATCAGGTGTACTTGACTCTAACAATAATGTCTCCAACAGTACCTTAGGCGGAAACAGTCTGCAGACGCACGATCATGATATTAAGATAACGGGCACAGGAAAACATTCTCACAAAAACAAAGTAACAGTCCCTTATTATATTCTGGCTTTCATCATAAAGCTTTAATATATATGAAAAATTGAAAATATAAATTATCCATTAATAATAAAGAGGATATTAGCATGACTTCGGAGCCAAATCTGTTAAACCGGATTACAATTACTATTGAAGCTAATAATCAACAAGTAGCTAGAAAAGTATTGCATGGCTCCTTGCTTAATCAAGCTAATATAAATAAATTATTTAATTCATACTTTAATGAATATGAAATTAATAGGGGTGTTTATTTAGAAACATTAATCCTGAATCTTGGTACGATAAATTTCCATGATTTTAATTCATTGTTTCCTACTCTCCTAAAAGCTGCATTGAATAAAGAATTCAGTCAATATCAGATAAACAACCATAGGGAAGAAATGCTATTTAATGAGACAATATCAAATCAAGCTACTGATAAGTCTTACATATTTGGCGATAACAAATTAATTGATGCAGAGAATTTCATTCACTTTTTATATCAAAAGCATTCCACATTAAATCTAGTAGAAGCAATGGGAAATAATGGTATTGAAAAATTAACAAATCAGTTAACACAAATAGAAAATAAATTTGCGTTATTATTGGCAAAAAGTTGTTTGTCTGAGGAAGGCTTAAAACGACTCTTGGCTATCAAACAACCCGATTTATTAATCGCTATCAATCGCAGATTATCTGAAAGAATAAATAGACCACAATATCAGGAGAAGCTTGTTTCCTGCGGACAACTGATATTTAGTGCTCTGGGATATATACAACAGTACAATATACAGGAAATTCCTAAACCGGATGAAAAAGTTATTGCACGCATAACAACTGAACTTAATAATAATGGTTTGCTTAATACAATACCTATTATTACACTATTTCGTCAGAGTGGGATTAACGATTCATCACTAAATGATTGGCTAAAGAAAATCTGGCAGGTGAGATCAATTTCACAGTTATGCAGAAAGTATCTTTCTGCTAAGGAATACCAATATCTGTCAGAACATTTTGTTTCAAAGAGCGTCGATAAAAATAGATATGATGAAGAGCCCGTAAATCAGAGCATATTATCAAGGTTGAATAATAATTCCATTAAAGAAGGAAATAATCACAGTCAACTCTGTACTCTCAGTAGACTATATTCTGAACCCGTTGTATTACCTGAACAAACCATTCTACGTCAGGTTAGTAATACAGTAGATCAGAGCATATTATCAAGGTTGAATAATGCCTCCATTAAAGAAGGAAATAACCAAAGTCAACTTCGCACTCTCAGTAGACTATATTCTGAGCCCGTTGCATTACCTGAACAAACCATTCCACGTCAGGTTAGTAATACAGGTATATTAATTCTATGGCCAATGCTACCTACACTATTTAACCAGCTTGGTCTACTTGAGAAAAAGAAATTTATCCATCGTCAGGCCCAGTTTAATGCCGTTGATTTTCTTGATTACCTGATTTGGGGAACCGAAGATGTGAAAGTGGAACGAAAGGTTTTGAATAATGTTCTATGTGGGTTAATGGCTGATGAAATTACTGAACCAATGCCTATTGAACCAGAAAAACAATGGATAATAATTCAATGGCTGGACGCTATTATCTCCCAACTTTCTGGCTGGAAAAAGTTAAGTCGTAATGACGTCCGTCAATTATTTCTACAACGACCAGGAGAATTACTGATCAATGAACAGGAAATTAAAATCACAATACAGCAACAACCATTTGATGCTCTGTTAACTGATTGGCCGTGGCCAATGAATATGGCTTGTTTTAGCTGGTTGAGTCAACCATTAACCATTACGTGGTTATAACCATTGACCACAATGACTTAGTCTGAGTAAAAAATATGAATATATCGCCTGTTTTTTATGATTCATTGAATCAGGATAACGACCGTGATCTATCGTTTTTATTTAGCGAACTGGAACGAATAGATCTCGCTCTTCAACACCATTTTTATTGTGTAGAAAGTCAGCGAAGTGAGCTCCTGGATGAGTTTCTGCTCACTGAGGCGGAAGTGGTGACCAGGCTGGATAAGCCACTTGGTAAACCTCATTGGATAAATGATGATTATCTGGCGATATCGCAAAAGGGCAATGTAAGCCTAATGGCAGCGTCCAGATTAATGGATCTGATCGAACGCTTTGAACTGACTGATTTTGAGCGCGATGTTTTACTATTAGGCTTATTGCCCCATTTTGATAGCCGCTATTATCGACTGTTTTCGCTGATTCAAGGGGGACAACAGGGTCGATTACCTTCTTTTGCGCTGGCATTGGAACTGTTTTGCCACTCGGCGCTGGAGAAACAGGTACAGCAAGCGAGTTTTCTGCACCGGGCACCTTTGATGGGTTGCCAGCTATTATCCATCGATACTAGTCAAAAAACGCTGGCCTGGCTCCAGACTCCCTTTATTACTGACAGCGGGGTATATCACTTTTTACTGGGGCATCACTACATTATGCCGGCTTTAGAACATTGTGCTGAGTGGTTAACACCGACAGGGATTGGCTGTTATCCTGAAGGATTAAAACAAGTACTGGGTAACGTATTGTTATCTGACAACGATAATATTAGACCGATTGTCTTATTACGGGGAATGGCCGGCAGTGCCAGAGCTTATACCATTACTAATATGATGGCTTCAGAAGGGAAGCAAACACTGCTGGTAGATATATCCAAACTTGCTGATAGCGATGAAAAAAACATTATTCTTCAGATAAAGCATATTTTGCGGGAAACCCGCATGCATGGAGCATGTTTATTATTACGGAATTTTTGCTTGTTAGTGGAACAGAATAAACAACTATTGGACTCCCTGTCAGAGTTATTGAATCAACCTGAATTAAGAATTGTTTGCCTGATTGAGCCTTATTCCCCATTGGTATGGCTGAAAAAGATACCGGTATTACTGATTGAGATGCCACTTTTAACGCCTGCGGAAAAAGCCAGATTGTTAATTGCCAGCTTACCGGATAATTGTTCCGAGGATATTGATACGATAACTTTAAGCCAGCGTTACACTTTTAACCCAGAAACCCTGCCATTGATTTTGCAAGAGGCCCAGCTTTATCAACAGCAGCGAGATCCGCTGGATATCTTGCAGCAATGCGATATACGCCAGGCATTAAATTTGCGTGCTCAACAAAATTTCGGTCAATTGGCACAGCGGATTATTCCTAAGCGCTCATTAAAGGATTTATTGGTATCCGATGAGATTGCTCAGCAGTTACGGGAAATACTCATAGCAATTAAGTATCGGGAACAGGTTCTGGCGGGAGGGTTTAAAGATAAAATTGCCTATGGCACTGGTATCAGCGCCCTGTTTTATGGTGATTCAGGCACTGGAAAAACCATGGCAGCAGAAGTGATTGCTGACCACATTGGCGTTGACTTAATAAAAGTGGATTTATCTACAGTAGTGAATAAATACATCGGTGAAACAGAAAAAAACTTATCCCGTATTTTCGATTTGGCGGAACAGGATGCAGGGGTATTATTCTTTGATGAAGCTGACGCACTGTTTGGTAAACGCAGTGAAACTAAAGATTCCCAGGACAGACATGCCAATATTGAAGTTTCTTACTTATTACAGCGCCTGGAGAATTACCCGGGTCTGGTCATTTTATCCACCAATAATCGTGGTCATTTAGACAGTGCTTTTAATCGTCGTTTTACTTTCATTACCCGTTTTACTTACCCGGATGAAAAAATCCGTAAAAAAATGTGGCAGGAAATTTGGCCTAGAAATATAAAAATATCGGAAGATATCGATTTTAACGAATTAGCTCAACGAACAAGCGTGACTGGCGCGAATATCCGCAATATTGCTTTATTGTCTTCATTCTTTGCTTCAGAGCAGGGGAATGATGAAGTCAGTAATGAAAATATTGAAATTGCATTGAAGCGTGAATTAGCTAAAGTCGGACGATTAACATTTTAAAAGTTATCACAATGAAAGTATTGAAATATTAAATAAATTTATTACCAAAAAGTTATCACGATATAATTTAAGAGAGGTTTTTTATGTTAAACACGCAAACTATTATTGATGTCAATAAGGCAATGGATGCCATGCTGCGCGCATATCTGAATCAAGATATTGCCATTCGTTTTGATCTACCTGAATTGGATACTATGCAATCTGATGCGATGGTAAGTATCTTTCTTTATGACATTCATGAAGATTTACAGCTTCGCTCGGCAGAATCAAGAGGGTTTGATGTTTATGCCGGGAGGTTATTGCCTGGTTGGGTAAATATTAAATGTAACTATCTGATTACCTATTGGGAAGCTTCTAAGCCAGCGACTGATGCCAGCAGTCCGGATAGCCAACCTGATAACCAGGCAATACAAGTGATGTCACAAGTATTAAATGCCTTGATTAATAATCGTCAATTGGCAGGTATTCCTGGTGCTTATACTCAGGTTGTACCGCCTAAAGAGAGTTTAAATAGCCTGGGGAATTTCTGGCAATCACTGGGTAATCGCCCACGGCTTTCTCTCAATTATTCAGTGACAGTACCTGTTAGCCTAAACGATGGTCAGGATAGCGCGACTCCGGTTACCGCGGTTTCTTCTACAGTGGAACAAACGGCATCGCTCAGTCAAGAAGTGGTTAGTCATGCTTTACGCGAATTACTCATTACGGAATTAGGAGGAGGAGAGGATAACCGGTTGGTACTGAGTAAAGTTGAATTATCCGCAGTGAAAGAGACGATGACTCAAGACAGTCCGGCTCAGATGATTATATTGTTGTCTGTTTCAGGCATTACACGACAGGAATATTTGAAGGAAATTGATAATATCTTTGATCGTTGGGTAAATAATGCTGAAGTTATTACCACTATTGATGATTGTGGGATTAGAATTGAAAGTATAACGAAAGATAATCTTGTAGGAATTTAA
SEQ ID NO:94(不对称光杆状菌菌株ATCC43949PVClopT操纵子，pvc1-pvc16；例
如，对应于GenBank登录号FM162591.1的序列的基因PAU02112至PAU02099)
CTTTTAACAGCGTTTCGGGGCTGGATATTACTTATGATGTCATCGAGTATAAAGATGGCACCGGTAATTATTATAAAATGCCGGGTCAACGTCAGTTAATCAATATTACACTGCGTAAAGGGGTATTCCCTGGCGACACTAAACTTTTTGATTGGCTTAATTCCATTCAGCTTAATCAGGTTGAGAAAAAAGATGTTTCAATTAGCTTGACCAACGAAGTTGGAACTGAAATTTTAATGACCTGGAGCGTAGCCAATGCATTCCCAACCTCATTAACATCTCCTTCTTTTGATGCCACCAGCAATGATATCGCTGTTCAAGAAATAAAACTGACTGCCGATCGAGTCACTATTCAGGCAGCTTAAAGCATCACGATGATTGATATATCAGACGGGACAAAATGATCCTCAAAATTTGGCACAACGGCTACCCGTCCAACTAAATTTACCCTCTTACAGTTCACGCAAAATATCGCACAATACAATTGGAGGCAATATGCCAACAACAACTTATCCCGGCGTTTATATTGAAGAAGACGCCTCACTGTCACTTTCCGTTCGCTCAAGTGCAACGGCGGTGCCCGTTTTTACCGTTGAAGATGACAGTCAACTTCATACTCCTACCAGAGTGAATAGTTGGTTAGAATATCTGACAAAAAAAGCAGATAAAAAATTCAATTCTACCGACAAACTTGATATCGCATTGCGCGCTTATTTTATTAACGGCGGCGGATATGGTTATCTCGTCAAAGCGGGTGAATTAACAAATCAAATTCCAAAACTTAACGATGTCACATTACTGGTCGCGGCTGGAGAAAATATCAAAGATGCTGTGAGTACACTTTGTCAACCGGGCAAAGGCTTATTTGCCATTCTGGATGGCCCAACCGAAGAGTTAAAGTCTGATGGCAAATCCAGAGATCCGTATGATCAAAGCCCTTTTGCCGCCGTTTATTACCCCTGGCTAGTTGCTGATTGGGCAGACAATATTCCGCCAAGCGCGGCCATTGCCGGTATCTATTGTTCAGTTGACCGTACCCGCGGTGTCTGGAAAGCCCCAGCAAATGTCATATTACAAGGCGGGGTGAAACCGAAGTTTAAAGTCACCGATGACTTACAAGGTATTTACAACACCGGTAAAGCCATCAATATGATCCGTGAATTTCCGAATACCGGTGTCACCATCTGGGGCGCCCGCACACTTAAGGACGAAGATAACTGGCGTTACATCCCAGTTCGCCGCCTGTTTAACAGTGCAGAGCGAGACATTAAAAATGCCATGAGTTTCGCGGTCTTTGAACCTAACAGCCAACCCACCTGGAAAGCTGTACACCGAGCTATTGATAATTATCTCTATGCCCTTTGGCAACAAGGAGGGCTAGCAGGAAACAAAGCTGAACAAGCTTACTTTGTGCAAATTGGTAAAGGGATAACCATGACCGATGATGATATCAAGCAAGGGAAAATGATTGTTAAAGTGGGTATGGCCGCAGTGCGCCCGGCTGAATTTATCATCCTTCAATTTTCACAAAATGTAGCACAGTAACCGTACTGAGGCGCGGTTTAACACCGCGTCCATTCAGTCTATTGAATGGAGGAGACAATAATGATAACGGAGATAAAACAGCCGGGCGTCACCATCACGGAAAATTCGATATCCCCGAAATCAGATAATGAATTTATCGGCGTCCCCGTTTTTATTGGCCATACCGAAAAAAATTCAAGCCATAAAACGGCTGTTAAACTAAATAGCCTGATGGACTTTACCCAAGCTTTCGGTGCATCAGGATTAACCTATTATTCAGTACGCCACTTTTTTGAAAATGGTGGACAGCAAGCTTATATCTTGTCACTGGGGATTAATCAACAGCTAAAAGATTTTCAATCATTGATTACCGCCCTGCAATGGAACTGGGTAAAACAAGCCATTGCCGCAGAAAACGAAATCACATTGATTGTTGTGCCTGATATTACCCGTTTTAATGATCTCAGCGCTCAAAAAAGCCTTTGGCTACAACTCTGGCAATCAATACTTGAACTGTGTAAAAGTCGGCGTGGCATCATGGGATTACTGGACGCGCCTGATGATCCAACATTAGCAACTGAGTGTTTAAAACAATTCTCTTCCACTGATCGCCAATGGGGCGCCGTATACTGGCCAAGGCTAAAAAGTACCTACCAAGAAAACGGTACATACATTGTACTTTCACCTACTGCTGCGGTCGCCGCCGTTATGCAACGCAATGACAGTCAGAAAGGCATATGGACTGCTCCCGCCAATGTGGCTTTAGCCAACGTCATCGGTCCGGTACGTTCTTACATTGAAGCTGGAACCTTGCTGAATCAAGAAGGCACTTCGTTGAATCTGGTGCGTAGCTTCCCCGGCAAAGGCATTAAAATCTGGGGCTGCCGCACTCTGGATAACATACCTCATTCTCCCTGGCGTTATATCCAAATTCGCCGTTTGGTTTCCTATATCGAAGCTCATATAACCCAACTTGGCCGCGCCTTTGTCTTTGAACCCAACAACGCCATCACCTGGATGAAATTTAAAGGTCAGGCCCACAACTGGCTACGTCAATTATGGCTAAAAGGTGGATTACGGGGCACTCAGGAAGATCAAGCATTTGAGGTGTTACTGGGTGTTAATGAATCCATGAGTGAAACGGATATCTTGGCCGGAAAAATGATCATGAAAATCAGGCTGGCGCTGTTAATTCCGGCAGAATTTATTGAGCTGAGTCTGACGTTTGATATCCGTAACAATACCGTACCTAGCTAATCTAAACAGGGGAAAAACATGTACAACTTATACACCCCGTCAGTATCTCACCGTTTTATCGCCAGTTTTCTGTTTAACAACATTCCCAGCCCACTTGATATCGCCTTTCAGCGTATATCTGGCCTGAGCCGAGAACTGCAAACCACCCAACATAGCCAAGGTGGAGAAAACGCCAGAAACGTCTGGTTATCCGAGAAGATCCAACATGGCAGCCTGGTGCTGGAGCGCGGTGTTATGACCATCACTCCCCTCACCTTGGTTTTTGATCGCGTGCTGCGCGGTGAAAAAGCCGTGTATGCCGATGTTGTCATCATGCTACTGAATGAAAATGCGTTACCCGTGGCGAGCTGGACAGTCAGTAACGCGCTACCGGTTCGTTGGTCCACCAGCGACTTTGATGCTAATAGCAACACCGTACTGGTGAGTTCTCTGGAATTACGTTATCAGGATATGCGCTGGTTAGGAGTAAAAGCATGACGGTAGAAATTAAAGAACTGATTATTCAGGCTAAAGTCACCGATTCTACGAGTGATCAACTCGCCCCAAGAACATTAGCCCAAGAAAAGCTGGATAACGCCCGTTTGATTGACATAGTGAAACGGGAAGTGTTAGAGGCATTACGTGAAGGAGGCCATCATGAGTTTAATTGAACGTGGTTTATCCAGACTCACCCTAACCGCTTTTAAAGACCGAGAAGGTAAAGTTTCCGTGGGTCGCTTACAAGCCATGTATAACCCCGATACGATCCAGCTTGACTACCAAACCCGCTACCAACAGGATGAAAGTGTTAATCGTGCCAGCCAAAGCAGCCGTTATGTATTATCCCAACCCGCCGGATTATCCTTAGTTCTGCTGTTTGATGCCTCGATGCCCGATAATAACATGCCGATAGAAACCCAGCTTGCGACCCTGAAATCCCTGTGTGCGATTGATGCCAGCACCAAAGTACCCCACTTCCTTAAAATCAAATGGGGCAAAATGCGCTGGGAAAACAAAGGTTATTTCGCCTGCCGAGCCAGTAGCCTGGCCGTCAACTATACCCTGTTTGACCGGGATGCCACACCATTGCGGGCCAGCGCCACTCTATCTCTGGTAGCGGACGAAAGCTTTATTATTCAAGCTACCGAACGGCAGTTAAAATCACCGCCGGCCACTGCGGTTAGCGTAACTGATATGCTCTCCCTGCCTTTGATTGCTTTAGATGCTGGAGCGTCTCTGGCTGGTGGCATTGATTATCTCTCGCTGGCCTGGCAAAACGGTCTGGATAATCTTGATGACTTTACCCCCGGACAAACACTGCAAGCGCGGGGGGATGCATGAAGATACCCATGATAACCCTCAAAATAGGTGGCAAAACGCTCAATCAATTGACTGTCATCAGTCTGACAATAAACCATCAAATCAATGGCATTCCCTCGACCAACATCACCTTGGGGATCGCTGGCGATGCGAGCCATATTTTCGACACCAAAGCCCAAGCTGAACTGGCAAGTTGTCGCCCCAATAATGAACTCACCCTACAGATCCAAAAAACCGTGGTGTTTAAAGGGAGCATCGTTCGACAAGCACTTGAACTGAAAGGTCAAGACAGCATCATTACCCTGACAGCAAAACATCCACTACAAAAGTTAACTCATAGCCTCCATTCACAATTATTCAGTCAACAGAGTGATGAAGCGATTATCAGGAAATTATTCAATCAGGCGGGTATCCAAACAACGATAAAGCAGGCTCCTCAACTTAAAACCGTTCATGAACAAATGGTGCAATTTCGTTGCAATGACTGGGCATTCCTAAAAAGCCGATTGATTGCCACTAATACCTGGCTGTTGCCCGGCAATGAATCGGTTACTTTGATAACACCTAAGGCCCTGAATCAATCGACAGTGCATACTCTTCATCGACAGGCCAGTGCTGAAGATATTGTGTTATTTGCAGCGGATCTCCAATGGAATAACCAATATAGCCCTAAAACGGTGAGTGTACGTGCCTGGGATATTGCTCAACAAAAGCTTTCCCCAGCAATTAATACCCAAAACAGTCAGCTTGGCAGTCATAAATTGGCCGTGGACAGTATCGCCGCACTGGCTGATAAAGAGTGGCAATGGGCTTACAGCTATCCATTAGATAATGAACAAGCCAAACACCTTGCTCAAGGCATTATGAATAACCTGCGAAGCCATAATATATCTGGCAGTTTTGAAATCGAAGGTAATCACCGTTATCAACCGGGGGATGTCTTGGCGTTAAATGGTTTTGGTCAGGGGATGGACGGTCAAGGGATTATCACCGGAGTCAGTCAGATAATTAATCAGCGGCAAGGCTGGCACACCCTATTAACCTTAGGCATGTTACCCGATGTAGAACCGCCGGTGCCTCAGGTGAAAGAGTTGCATATCGGTATCGTGGAAAAATACCAGCAAGACCGCCAATCACTAAGCCGTATCCCAGTCAGAATACCCGCATTAAACTTGACCAAAGGTGTCCTTTTTGCCCGGCTAGGTAAACCTTATGCCAGTCATGAAAGCGGATTTTGCTTTTATCCCGAACCGGGAGATGAAGTGATTATCGGATTCTTTGAATGTGATCCTCGTTTTCCAGTGATATTAGGTTCCATGCATAATCCGAAAAATAAACCACCGTTAGAACCCAGTGAAAAAAATCCGGTGAAAACTTTAGTTATCAAGCAAGGGGATAAACAACAAGCATTAATATTCGATAATAAAGAAAACACGGTGGCACTTAATAGCGGCGAAAATAAAGTCTCTCTGCAACAGGATAAAAACATTACGCTCAATTCAACTAAAAATCTCATCACTCAGGCCCAAGAAATTAATATACAAGCGGAAAAATCTCTGTCAGCCACAGGAAAATCTGGCGTCGATATTAAGGGCGCGAAAATTAACTTAACCCAGTAATGAGGTATTGAAATGACAAGCCAAATATTAGCCAATATTTACGGTTGCGGCTGGAAATTTCCGCCACAGTTTTCTATTGAAACTGGCGTAGAAATGGCCGAAGGTGCCGAAAACGTTCGCCAAAGTATGAAAATCCTTTTTTTAACTGAACCCGGTGAACGAATTATGCGTGAAGATTATGGTTGTGGTCTGAATGATTACATGTTTGAAAATATCAGTGATGAATTATTATCGGAGATTCAAACCCGCATTGAAGAACGAGTATTGCGCTATGAACCCCGTGCTGAAATCACAGATATCCAAGTAACTCAGAAAACAGACTCACCGAATACTTTACATATTCAAGTGACCTATGCCCTGAGAGGCAGCCAAATCAGTCAACAGCTTGAAGGGGTTCTTGAGATCAACGAAGGTCAGGCAAAGGTGAGTCTATGAGCAAACAACTCATTATTGATGGCGACAGCCTGCTATTCGAGCCATTATTCGGCAACCGGCAGGTCACTATTTTGATGCCAGCGACCATCAGAGGCAGCGGACACGCGCAAATCCAAGGCAGAAAGATAGCGATTGTCGGCGATGAAAAAAAGGTACAACTTCAAGCGCAATACATTACCCCAAGCCACCCGGTACCTGGCATAGGCACAGTTACCATTGCTCAATTAGATACCAGCCAGCAAGTCAACTTTTGCCACAGCCCTGCCACAGTGATAGTTGTCGGGCAGCAATTTACCGCTCGATTTACCCCATCACAGCCGGCAATTAATCCGTCAACCGGGCCAGATGTCACAACACCCAGTATGGGCAAAGGCCGTTTTATTGCCAGTCAACATACTATCAACGCCGGATAAATAACTCTGCAAAATCATTATTCAATAACGTTCCTATTCTGCAATAGCTATCAGCAATATATTCAAATAACAGGTGGTATAATATGGGACTCACCGAATTAAAAAATAAACTCTCTGCTATCGTACTCGATACGGATTTTAAACTTGATGAAAGAAGTACACTGGATATTTTAAACTGGCTACAAGAATATGCTAAAAAAATCCCTTTCAATCAAGAGAAAAAACAGTTCTGGGATAGTTTCTATTTTATTCAGGAAAATAGTCCTGAGAAATTAGCCGATCTTTACCAAAACGTTAATAAAACGAATGGCCATTTACCGGCCCATCAAGCTTTTGTTTTAGCCTTTTTAAAACTTTTAGAAACCACCAAAGTATTATTTAATACTTTTCCGGCACGACATCGTGATCTTTATTACCGGGAATTATTAGGTCTAAAACCCAGAAATGCCCAAGCAGATAGTGTTGCTTTAGGCATTACCTTAAATACAGATAACACAGAACATCTTATTCCTAAAGGAACCTTGTTCGATGCCGGGCAGGACAGGGCCGGAAATCCGCTACAATACGCATCAAATGCAGATTTACTGGCGAATCAAGGAAAATTGAGCGATCTGCGTTGGTGTCGAAAAGATAATGATAGCTGGCAATCTGCAATACTACTGAACCACTCAGATAATATTGAATTACCTGAAAACAGTATTCGACTTTTTAGTCCAACGCCGGATGATATTCCCGTTTTATCCGGTTATTTGATAACTTCGTCTTTATTTGCTATGCCAACGGGGGAACGCAGTATTACATTGACTTTAGCAGATAATTGGCATGGTGATATTAAGCACATCACCGCTAAAATCAGTTCGGGAGATCACTGGCTTTCACTATCAGTAAAAAAAGAACAAGACAATAGTATTCACTATCTTAAACTTTATTTATCAACCAATGATGACCCCATCGGTCCTCCTGATGCTTTGGATAATATAGCGTTTGATGTACCGGTATTAAAGCTGGGCACTGTTCAGGGACCTATACTACCCAAGATTACGGGTATTGAAATTAGCATTAACGGCAACAGTAATGTACATTATTCCTCTGATAACGGTATTGAAAAAATAGATGCAGCTAGTTTTCCCTTTGGACAATCACCGTCACCAGGTTCCGGTTTTAATCTGATTGCCCCTGAATGGTATGGTACAGAAAGCGCCAAAATTACTCTTACTCCTCAATGGACTGGATTACCCAAAGAGGGGTTTAAAGAGTGGTATCAAGGATATAGTTCTACCCCCGAAAATAATGCATTTAAAGTACAGGCTTATTTAATCACACCTCAAAAGAGAGAAAAATTTAATGAAGCTCAGTCATTATTTAATGAAAGTAAAGACAAGAAACCACAAGGAAAAAGCCTAACTTTTACCTTACCTGCAATGGATTATTCCTTTGCAAACAGCCCATCATCTAATAACTGGCCCGCATCAATACGCATAGAACTAACCGAACAGGATTTTATGCATGCCCAATATTGGCAAAATCCTACGGGTAAAAAACAGCCCTATACCCCCAAAATGAACACATTACAAATTCAGTTCAGTGCCAAAGTTAAACCCGAACAATTTTCCGTTTATTCTCTCACGCCTTTTGGTTGGGGAAAAACAGGAGAAAATAGAACATCATTAACCCATGATACATTCTATTTAGGTTTTACCGATGTATTACCAGGACAAACTTTATCCCTGTACTGGCAGTTAGAAGGTATTAAAAAGCTCCCTTTATCCTGGTCTTATCTGAATCAAGAAAATACCTGGAGTCCATTGGATAATCAGGTGCATGACCAAACCCACAACCTATTTGATCGAGGAATCTGGCGTACCTCATTGCCACATGATGCTTCAAACCAAGCCTCTCAAATGCCAAAAGGACAATATTGGGTGAAGGCACACATTTTACAAACGAATCAAGCAACCCTGACTGATCTGTATTGGTATCGAAAAGATAATGATGTCTGGAAATCCGCAACACCTCTTAGCCTTTCAAATAACATGAAATTACCCGCAAACGGTATTCAGATTTTTAGCCCAACATCTCATGATGTTCCAGTTCGATACGGCTACCTAATTACTTCATCTTTATTCTCATTCCTCAAGAAAGGACGCAATATCACATTAATTTTAGCAGGAGATAGCTGGGAGGGTAATCCTGAAAACATCACCGCTAAAATCAGTTCAGGAAATCACTGGTTAACACTATCCGTCGAATATCTGAGTAATACTAATAGTCTTAAGTTGCAATTATCAGATAATAATAATGATCCCATCAGCCCCCCTAATGCTCTGGATAATATGACGTTTGACACGCCATTGTTAAAACTAGAAGCCACTCAGGATTTCACTTTGCCCTGGATTTATAAGGTATGCGTTAATAGCAACAATATACTCTCTACCTCTGACAGCTCAGATGCAGCGATTACTCGTTTCCCCTTTGGCCAATCACCATCGTTGGGTTCCAGCTTTAGTCCGAAAATCGTTTTCCCGGAATGGTTTGAATCTGAATACGCATCAGACACCACGATCACGATTACCCCTCAATGGGTTAACCTGCCCACAGAAAACTTTTCATCGTGGTATGACGGATATATTAATAAACCTGCCGATAATAGCGTATTTAAAATAGAGGGTTATTTACTTACTCATTATCAGGGAAAAATCAAACTCACAGAAGCTGAGACAGGAAGCGAAACCCAAGCATTATTCAATGGAAACAATGCACCACAAGGAAAAAGCCTGACTTTCACTTTACCTAATAGGTATAACTTCTATCCGCGCAACCATCAGTCAATGAAGATAGAAATAAAACTCGTTAAACAAGACTTTATGCACACTCAACATAAGAGCAATCCCACAGGCAAAAAACCACCCTATACCCCGCAAATCAGTGCCTTACAGGTGGAATTCAATGCTACAGCTTTCCATCGAAAATTCTCCGTTTATCCTCTCACGCCTTTTGGCTGGGGCAAAACAGGAGAAAATAGCACACCATTAATTCATGATACATTTTATTTAGGCTTGACCGATATATCACCAGAGCAAACTTTTTCTCTGTATTGGCAGCTAAAGGGCCTTAAAGAGCTACCTTTGTCTTGGTTTTATCTAAGTGAAGAAAATAGCTGGAAATCATTAAATAGATCAACTTACAACCAAACCCACAACCTGTTTGAATCAGCAGAACAAAGTATCCTATTACCACGGGATGCTTCAAACCAAGCCTCTCAAATGCCATTAGGACGGTATTGGCTGAAAGCACAGATAGAACAGGAGAAAAAACAGATAAAGATAGCGCTTCCTGATTATTATCCAAGAATCAGGGGGCTGTTGTATAACGCTACCATCGCCACTTTAATCAACGCTGAAGCTGTTGAGCAATCTCACCTTATCAACGGATTGGCTGCTAACAACATTAAACAACCGGTTAACTCATCCGTTGCCATCAACGAAGTTATTCAACCCTGGACATCCTGGAACGGTCGCCCAAAAGAAACCGAGTCAGCATTCCTGGCACGAGTTCCTGCCCGGCTCTCTCATCGTAACCGAGTGCTAAGCTGGGGTAACATTGCCACTTTATTAAAAGAGAATTTTAGTAGCTTATTCGATGTCAAATACCCTTCTGTCAGTGAATTAACCAAAATTCCAGCGCCAGAAAAGCGACAATTAACCATCATCCCCGACAACCGCTATAAAGATAATGATGATTCACTACGCCCAGTATTGAACCAAGCCAGACTGACCGAGATGGTCGAATGGTTAGATCGATTAAGTAGCCCTTGGACAACTATTGAAATTAAAAATCCCACATATGTTAACGTTCTGATCCACTATGAACTGATATTTACCTCGGATGTTAACCCCGATTATGGCCTCCATCAGCTACAACAAGAACTCAGTCGAAAATATATGCCGTGGGGAGAAAATGCAGCTATTGGCGTAACACCCGGTAATCGTATTGACTACTTCCAGTTATTAGCCTCAATTCAACAATCACCGCTGGTTGAACGGGTCACCAACTTAACGTTAAAAAAAGGCAGCCAGCCTACCGTAAGTGAAAGTATAGAAGCCGCCGATGATGAAGTACTGATTTTAGTCTGGTCATAAAAACTTCCCCAACCTAAGGAATTAACAAATGAATAATCGAGATATGCTATTTCCTATCATTAAAGACGATATTACCTTTGATTCTTTATTCGCCCAGGCAAAAGCCGTTATTGAACAACAATCGGGGCAGCTCTGGAATAATACAGGTGAAAATGATCCCGGCATTACTTTATTAGAAGCCTGTTGTTATGGCGCATCCGATCTGGCCTATCGCCACACATTGCCACTGCGAGATTTGCTTACTCCTCAAGAAAATGAACGAATAGATGATGGCATTTTTCCCAAAGAATTTGGTCCACAACAAATACTGACCTGCGGCCCAATTACCGCGGAAGATTACCGTCGAGCTTTGTTAGATTTGCGTAGTGATAACACCGTTGAAGGTTATTTTTTCTTTAATGATGCACAGCTCATTCGTGAACCGGAAAATCAACGCTATTCATATTGGTATAACAAAGAAAAACGCGAATACAGTTTTACTCAAGACCAATACAGCGAACAATTACAGTTAACACTGAGAGGAAACTATTGGCTCTATTTACTTCCCAGTCGGAAAACCCAGCTCGATAACACCCTGGCTGAAGAAAGACTCAACATTTTTCTGAAAGATAACCGAAACTTAGGAGAATCGGTCAGTAAAATTATTTGGCTAGAACCCATTAAACTGTCATTGAAAATTGATATTCAGCTTGATGATGACGCCAAAGATATTGCTGATATATTTGCTAAAGTTTATATGATTGCAGAACAAATGGTGCTTGAAAAACCATTACGTTATACCACTCAAGCGATGAAAGAACTGGGTTACAGTCAGGAACAAATATTTGAAGGCCCTTATTTACACCACGGTTGGATACCGAAATTACCTCAAACCAAAGATTATACTCACCCTACCGTATTAAATCTCAGTCCTTTAATTAATCAGTTACTGGCTATCAAAGGGGTGAAACATATTACCCAATTTACATTGGATAAGCCTGATAAAAAAATTTCTAAGTTACCAAATGATAATTGGTCTTGGGAAATCGCTCCGGGATATTACCCAAAACTATGGGGAGATACTCCATTAGAATTAATTACCTCACCAACAAGCCCACTCACCATCACGGCAAAAGGGGGAATTAAAATTGCTATTACTAAACAACAGATAGAAAAAAACATAATGACAGAACCACTAATTAATACACAGCCAGAATTATTGAACTGGGGTAAACATCGCAAAGTCCTGGATTACTATCCGATAAGCAATAAATTACCCGCTTGCTATGGATTACAAACTAATACCCAACAACAGCTACAGTTGCATCAATTTATGCTGCCTTTTGAACAAATGCTAGCGAATAACTGCGCTGAACTTGCTTTATTGCCAAGACTATTAGCTTTTAAACAACGAGGAAATACGGTACATGGCATTCAATGGCCTTTTAAAGAAAATACGGTTGGTCAACATGTTCATAAGGACATAGTATCTAATTTAAACAATAATGCTACGAAAATCGATAATAATGCCGATGACTACGACAAGGAACTCGTTATTCTAGATTATTTGTTAAGATATTTTGGGGCTCAATGTGCAATCCCACGACTATCACCAGACCCACCACAATCATCATTAACAGAACCTCAGACTAAAAAAGATTTTCTATCTACTCAGCGCGAATATCTGGCTCAACAGCCAAAACTGACTTATCAGCGTAACAATATTCGGATTGATAAAGTATCAGCACTGCAAAAACGTATCGCTGCCCGATTAGGTCTGGGAGGAGAATGTTTCAAAGCAGAGCCTGACTTAGCTCACCTTCCTTTCTACCTCATTGAACATCGTAGGCTCTTACCAGTAAAACCTGATATAAAATTCTATATTGAGCAACAACCTAATTCTCTGGAAATTGAAAATGATAAATTAAAAATCACACAGAAAGATTCAGCGGGTCGGTTACTGCAAGGTCAAGTTATTAACCTGGAATTTCGTGAGGGCTATGATGAATTTACATTGCTAAACTTAATGATAACTGAAGTGACAAGAGATACATTCACCATTAGCATTAATAATAGCCGTGATCTCAGAGACAATCTGGACAAAGTGCAACACGCGTTTGAACAAACGAATAATCTGAGCTGGCACAATAGCTTAATATGGATGGAAGATATGGATTATCAATTGGTTTATGCCAATGGAGAACAACTGGAAAAAGCGGAAAATGAACGATGGATTACCATTAACAATCAAAGTGCTTTCCCTGCTATGATCGGAGAGAATGATGAAATCACACTAAAAATTCAATCCGATTATGAACTTAAAACCAAAGTCGTGCGGCTTGATTATAACAACAAAAAAATTCTGATTATAAAAGATGCGACATCAATAAATAATTTTCCGCCAAAAAGAGAAGCATCATATTATTCTTGCTCTTCTCTAAAAGACAATGGGTACGGATATTCGGATGAATATAAATATGAACTTACTTATATTGATACAGATTCTACAAAAGAAAATGAGTGCTGGATTACTATCAGCGATCCAAATAATTTGTTTTCTCCTGATATCATCGCAGAGAATGACGAAATTATATTGAAAGCTAACCCTAATTATGAGTTTAAAACGCACGTAGTAAAATTTGATCGTATTAATAGACAAATATTACTTAGGAAAAATACAGACCTGGAAAATAATTTTCCATCAGAAAACAACACATCGCACTATCGCTGGCATTTCTCTGGTGAAAAATATGCCCAAACTGACCATTTTTCATTTGTTGTCAGTGCAGTACTGAATCGAGAATTAATTGAGAGGGGCACAGTCGATCTCTATAAATTAGAGTCTTGGGTAAAAACTGAGATTTTATCTGAATTACCCGCGCATATCTCACTCGTTATTCATTGGCTATCATCGGAAGAATTCGAAAAATTTGCCAGTACTTATAAAGTTTGGCAAAATAATGGCGCTCCTTTAGGTGATCACGCATATAAAATTCTAGAAACATTAACACTTGGGAAAAAACCTTCTACTTCAGCAAGAAGGTCCAGCAGCTATATAGAAGCACAGTAATAATTCTTACAGAACATTAACCCATATTTATCTTATAATATCAAACATCATAAAAACAATCTTCAGCTCATTATAATGACATATTTCATACTCAGGTTTCTTCATATCTGTTAATTACAAAGAGAATATTAATATGATCTCAGCACCAAATCTGTTAAATCGGATTATCATTACTATTGAAGCGAATAACGCACAGGCAGCTAAAAAAGTATTGCATGGCTCCCTGCTTAATCAATCCAGTATAAACAAACTCTTTGATTCATACTTTAACCAATATGTTGTTAATCAGACTATCTACCTGAAGACACTCACCCTGAATCTTGGCGAAATACGATTAAATAGTTTTAATTCACAGTTTGTTATTCGGCTTAATACTATTCTGAGTCAAGCATTGAGCCAATATCAGGTAAATAATCAAACTGATATTGAGAAATTTATTTATTACTTATATCGAAAAGATTCTATATTAAACCCAATAGAGGAAATCAATAATCGTGAAATTACTGACATCAATATTAAGCAATTAATTAACCAATTACCCCAGATACAAAACAATTGGACACTATTATTGGCAAAAAGCTGTTTATCCACACATAGCCTGAAAAAACTCCTGGCTATCAAAAAAACAGCTTTATTAACCGCCATTAATCGTAAATTATCTGAAAAGATCAATATATCACCCTATCAGCAGGAATCGGTTTCCACCTGGCAATTGATACTGAATGCGCTGAAATATATACAGCGACATAATACACAGGAAATACCTGAACCCGATGCGAAAGTCATATCACTCATTACAACGGAACTCAATGACAATGCCATTAATACAGCACCAATTATTGCATTATTTCGCCAAGTTATAACCAACCATTCCCCACTGAATAAGTGGCTGGAACAACTGTGGCAAACAAAGCGAATTTCACAGTTATGTAAAAAACAGCTGTCAATTGAAGAATACCAACATCTATCGGAGCGCTTTATTGCCAAACACGGGAATAAAAATAAATCTGATAAAAAATCATCCATGACTTCCGAACCGCTGTTATTACCTGAACACCCTCCACCACGTCAGGTCAATAATGCTGGAATATTAGTTCTGTGGCCGATGTTACCTACTCTATTTAACCAATTCGGCCTGTTTGAAAAACAAAAATTTATTCATCGTCAAGCTCAATTTAGGGCTGTTAATCTACTTGATTATCTCATTTGGGGAAACGAAGAAACACAGACAGAACGAAAAATATTGAATTGCGTTCTGTGTGGGTTAATTGCCGATGAGGACACGGAATCAATCCCTATTGAGCCAGAAAAACAACAGGTAATAGAACAATGGTTAGATGCAGTTATCAGTCAACTTCCTGCCTGGAAAAAATTAAGCCGCAATGATAGCCGCCAATTGTTTTTACAACGCCCGGGGGAATTGCTGACAAATGAGCAGGAAATCAAAATTACGGTACAACCTCAACCATTCGATGCACTGTTAAATAACTGGCCCTGGCCGTTAAATATCGCCAAACTTCCCTGGCTGGATCGCCCTTTATTAATCAACTGGTAAAACATTGACAAGGTTTATATGAAAGAACATCAATATAGAATAGTCGATCTACGCTGGATTTATTCCCATTTGGAGCGCATCGATCTGCTGTTACAACGTCACTATTACCAAAAGAGAGACAAATACGATTCATTGCCAGAAAGTTTTTTGCTTGAAGAAGATGAATTAGAACAACGTCTAGCAAAACCGTTGGGTATTCCTCATTGGCTAACAGCAAATACCGGCGCTGGTGATACAGAAACAGAAAATCATTCTGCTTCCGGCACATTATCACTGCTAGTCACGCGTTTTAAACTCACTGAATTTGAACGTGATGTGTTATTGCTAGGTTTATTACCGCATTTTGACAACCGCTATCATGCGTTATTTGCTACTCTGCACGGTAACAGTAAAAAACAGTGGCCCAGTTTTGATTTAGCGATTGAATTATTTAGCCAACATCAAAGTAACTGGCAATTATTTCAACACCACTTTTTACCGCAAGCTCCATTAATCAATCACCATTTATTACGACTCAATAACCAAGAGGAACCCATTTGGCTACAAACTCAATTTTTAACTCACAATGCAGTCTGGTCTTTTTTATCCGGTCAGCGCGTCATTTTACCTCCCTTAATATCCTGCGCTTACTGGCATATTCCAACCTCACAGACTTGGTATCCACCAATCCTTGGTCATGCATTTGAAAAAATATTGCTGAATGAAACGGACGAAATACGCCCGCTGGTGGTTCTTAAAGGAAAACAGGACAGCGCCAGAGAACTGGCAGTCAGTAATATTATGGGAATTCACGGCATTAACACTTTAACGTTCGATTTATTTCACCTGCCAGATGAAGAGTGCACCACCTCAATACTCAATCTGCTAATAGATGCAATACGAGAAACCCGGCTACATAATGCCTGTTTATTAATCCGTAACTTTTCTTTGCTGGCAGAGGAAAAGAGAATATCGCATAGAGAATTATCAGCTCTACTGAATCAACCCAAATTACGTGTGGTTTGTCTGGCAGAGTCAGAAGAATCATTAGCATGGGTTAAACACCTGCCGATAGTGCAAATTAATATGCCACCGGCGACGCTGGCAGATAAAAAAACGATGCTGGAAGCCAGTTTGCCAGATAATGTCACTAAAGGAATTAATATAACTCAATTATGTCAACGTTTTTCATTTACAGCAGAAACATTACCGTTAATTATCAAGGAAGCTCATCAATACCAAATCCTCCGACAACCGGAAGATCAATTGAAAGAATCTGATCTACGTAAGGCATTAAATTGCCGCGCCCAACAAAATTTCGGTAAATTAGCCCAGCGTATGACACCAAAACGAAGTTTTAATGATTTGGTTATTTCCGCTGACTTAACTCAACAGTTGAAAGAAATCATCGCAGCAATTAATTACCGTGACCAAATTCTGGGCGCAGGTTTTCGGGAAAAAATCAGCTATGGTACTGGTATTAGCGCCCTATTTTACGGTGAATCCGGGACGGGGAAAACCATGGCCGCAGAAGTGATTGCCAGCTATCTTGGTGTTGATCTGATTAAGGTAGATCTTTCTACCGTGGTGAATAAATACATCGGTGAAACCGAAAAAAATATCTCCCGTATTTTCGATCTGGCCGAAGCGGATTCCGGGGTGCTGTTTTTCGATGAAGCCGATGCCTTATTCGGTAAACGCAGTGAAACCAAAGATGCCCAAGATAGACATGCCAATATTGAAGTTTCTTATTTATTACAGCGACTAGAAAATTATCCGGGATTAGTGATTTTAGCGACTAACAATCGCAACCATTTGGATAGTGCGTTTAATCGCCGCTTTACCTTTATTACCCGCTTTACTTATCCCGATGAAGCATTACGCAAAGCAATGTGGCAGGCAATTTGGCCTGAACAACTTAAGTTATCAGATCAACTTGATTTTGAGCATTTGGCTAAACAGGCAAATCTGACCGGTGCTAATATCAGAAATATTGCCTTATTATCATCAATATTAGCTACAGATAATAATAGTGATCAAATTGAAAATAAACATATAGCGCGAGCATTGATACTTGAATTAAATAAAACGGGCCGATTGATTTTTTAATCATTTATACCCAATAAATTTCGAGTTGCAGCGCGGCGGCAAGTGAACGAATCCCCAGGAGCATAGATAACTATGTGACTGGGGTGAGTGAAAGCAGCCAACAAAGCAGCAACTTGAAGGATGAAGGGTATATAGAATTGGAGTGAATATGACAAATATAATTAACCCTAATAATGCGATTCTTGAAGTTAATAACGCATTAAATGATATTTTATCTCAGTATTTAACTAATATTGATATCCGCTTTGATCTACCAGAAATAAATTCAATCCCATCAACCCCTACAGTGAGTATATTTCTTTATGATATACATGAAGACCTACAATTACGTTCTGCTGAACCAAGAAGTTATCATCCTACCACCAGCTCATTATTGCCGGGATGGGTAAATATTAATTATAACTATTTAATTACTTACTGGCATTCAAGTAATCCATCAAGCGACAGTTCTACCCCTGATAGTCAACCCAATAATCAAGCGGCACAAGTCATGACTGCTATTTTAAATGCATTGGTTAACAACCGACAATTACCTAAAATTCCTGGCGCATATACCAGAGTCATTCCACCTCAAGAAAATCTAAATAGCTTAGGTAACTTTTGGCAAGCGCTTGGCAATCGCCCTCGCCTTTCTTTATTATATTCAATTACCGCACCGGTAAAACTGCAAAATATTAAAGATGTCATAAAGCCCATTAGCCAAATTTCCACTTCTGTGGATCAAAAATCAAATCTGGATAATTCGCAAATCAACCAAGCCTTATTTAGCAAATTGGGTGCCGATTTAGGTGGCACACAAGATGTTCGTCTTGCTCTTGCGAAAGTGAATCTGACAACCAAACCTGCTAAAGAAAATAATGAAAATCAAAATAATAAAAATGTAATTATTGAAGTTTCTGGCATTACCCATTTGGATTATTTACCCAGAATAAAAGGTATTCTTTCAACATGGGTAAATAGTCATAGTGCTGTTGTTAGGATAAATGATATTGGTATTATTGTTTCAGAATATAAATATGATAAATTAACAGGCGTTTAA
SEQ ID NO:95(不对称光杆状菌菌株ATCC43949PVCPaTox操纵子，pvc1-pvc16)
ATGAATACAGCTCAAGAAATTATTAACCGTTTATCGGGGAGAGCCGTTACGCTTGGTTGGGATGTTGTTATTGCTTATGACCGAAAAAAAATTAACACTCTGTTAGAGCAACAATATGTTGAAAAGGTAAAAAACGGGGAGAACTTCCCGCTTATCAACTGGGAGAACCAGAGAAAAACACTTCAATTTAAAGATCTTCAATTAGGTGTTCCACTTATTTCTTTTGAGAATTCAACACTGGAAAATTCAAGGGCGCTTGCCACGATAGAATTTATTTCAGGAGCTATTATTGAATTTAGTGACTCCGGGCAAATAATCAACTATAAGAAGATTGAACCTAGTCATGGTTATGGCATGGTGCTGACTATCGATCTCATGGCTGGTACAGGTTCAGTAGAAGAACAAGGTCGGGTGATAATAAATCTTAACGAAGGCGCCATACTCGATTTGCATGTTATCCAACAACCGCCAGCAGAAGTGGTAGAATTTTTCCGCACTTGGTTGATGGCTAATAAAATGACTTATGAATTAGGTAAGCTGGATCTGAGTAGTCAAGCTGGTCTAGTGCCTCGTTCTTTTCGTATTCGTACTCAGCGGGCGCCTGAAAAAATTCGTAAAGCGACGAGCGATGAAGGAAATGGCGCTGTTTTGTTGTTTGTTGCCACTAACTATAACCCTACAAGTGGAACTTTACCTGCCAAGGATTATCCGTGGCTAATCCCTGAGGAATATTCAGGCGCATTGCTTATCGGTAATAAATGCTTATTTAAAGACATTCTGAAACCGAATCTGGATCAGTTGTTTGATAAAGGGGAATGGACATTAAAAGTTCAGCAAACGGATTCTGATCAACTGCTGCATTATCTGGAGGCAAACTCTGCATATATAACAGATAAGCCTTATATGGCAGACTTTGAAGGAACTCAGGATGGAGTCTGGACAGGACGTTATAAATTTGAGACTGGCCGGGGACATTATGGGGTGTATGAAAATGTACGCTTTCCTATCAATGGAATGTTGATGAAACCGGCTAAAACTGGATTACAGTTATCAATAGATTCACCACAAAGCCATCAATTTAATGTTGATTTCGGAATGAAGTGGTTCCATTGTGCTAATATAATGTGTGGTTATTCCTGGTTTAACGAGACTTACCCATTTTATCTTGATGGAAAATCATTTTATCAAGTTCATATTGACCCTGATAAAGAGGTGATTTATTTTACTGGGCCAGATGAAGATATTAATATTGTAGGAAATTACAGCCCGCCTGCGTGGTGGCAATCTAAATGGCAAAAACATATCAGTGATGATTTTACGGATATTTCCTCGGAAAAATTTAAGCGACTCAGTCAAATAAAATTGCCAGAAATATGCATGTTTGCCGTGAACCATTTATTATTTCCTGGTCATAATACTTTGCTGTTGAAAGACGTTTATTTACCGGGTGATATGGTGATTTTCGGTGATATTAACCCATCACTTACCGCTTTTCGGGTTACGCCATTAAAAGCAACAGTGGTGGCAAAGGGAACCCAACAATTTAAAGCCATAGAAACTAATTGATGATTATACCCTTCATCCTTCAAGTTGCTGCTTTGTTGGCTACGTTCACTCACCCCAGTCACATAGTTAGCTATGCTCCCGGGGATTCGCTCCCTGGCCGTCGCGATGCATCTTGAAATCCATAGGGTATATATTTAATTGGATAAGTCTTTTTTATTTTAACATTATAACCTGATTCTTTTTGGATAAAATTAAAGGATTATTAACATGTCTATTACACAAGAACAAATCGCTGCTGAATATCCTATTCCTAGTTACCGTTTTATGGTTTCTATAGGAGATGTGCAAGTCCCTTTTAATAGTGTTTCGGGATTAGATAGGAAATATGAGGTTATTGAATATAAAGATGGCATTGGTAATTATTATAAAATGCCAGGACAAATACAGAGGGTTGATATTACACTTCGGAAAGGCATATTCTCTGGGAAAAATGATTTATTTAATTGGATTAATTCCATTGAACTCAATCGGGTAGAAAAAAAGGATATTACAATTAGTTTAACTAATGATACTGGCAGTAAAGTCTTAATGAGTTGGGTTGTTTCGAACGCCTTTCCGAGCTCACTGACGGCCCCTTCATTTGATGCTTCAAGTAATGAAATTGCAGTACAAGAAATTTCATTAGTTGCTGATCGGGTAACAATTCAGGTTCCCTGATAACTAAAAACTTTAAGGAAAAATAATGTCTGTACAAACAACTTATCCCGGAATTTATATTGAAGAAGATGCATCATTGTCTCTATCTATCAATAATAGTCCAACAGCAATCCCTGTTTTTATCGGTAAATTTTACAACTTGGATGGTTCCTTACCTAAAGTGGGAACATGTTCTAGAATTACCAGTTGGTTAGATTTCACTAAAAAATTTTCGGTAGCTCCTCCTCAAACCATTTCATTGATCGCGTCGCCAATTGCTGACACACAAGAAAGTGTACCCAAAGCAGTTCAATATACTTATAAGGCCGAGTTTGAAACCTCAGAAAATCTGGCAAATGGTGCCTATGCGGTACAACATTATTTCCAGAATGGCGGTGGTATTTGCTATATCATACCTTTAGTTAGCGTGAAAAAAGAGGATGCTGCGATTGAGTTAACAAAATTACCTGAATTAATTGAAAGACAACAAGAGATTACGTTAATCGTCTGCCCGGAGGACGATAAGACGCTCACTGTTGATAGCAGTAAAAAATCGGATGTTTATAACAGCATCAATACATTATTGAGTAATAAGGTAGGTTATTTTCTCATTGCAGATTCAGATGATGGCAAAGCAGTTCCTGATACGTTGCCGGAAAAAACTGCGGTCTATTATCCTGGTTTACTAACTTCTTTTACACAACGCTATGCCCGACCTGCCGATTCTGCTATCAAAGTGACCGGTATTACAAATATATCAACTCTGGCTGATATTCACACCAACTTGGCCGATGACTACTCAACAGCAAGTCAGGTTATTAATGATGTTTTGGAAAAAAATAATAAGCTCGCATCGTCTCCCATTATTTTACCTCCCAGCGCCGCTGTTGCTGGTGCTTATGCCGCTGTTGATGTGAGTCGTGGTGTTTGGAAAGCACCTGCGAATGTGATGTTAAGTAATGCCACGCCAATCATTAGTATTTCCGATGCGGAACAAGGTGTGATGAACCCATTAGGTATTAATGCTATTCGTAGTTTTACTGGTAGAGGTACTTTGATTTGGGGAGCTCGTACTCTGGATAAAACGGATAACTGGCGCTATGTTCCTGTACGTCGTTTATTCAATAGCGCAGAGCGAGATATTAAGTTAGCAATGCGTTTTGCAGTTTTTGAGCCTAACTCCCAACCAATTTGGGAAAAGGTCAAGGCTGCTATCAATAGCTATTTGCAGTCACTTTGGCAGCAAGGTGCACTGCAAGGCAATAAACCCGATGAAGCCTGGTTTGTACAAATTGGTAAAGGCGTGACCATGACAGATGATGATATTAAGAATGGGAGAATGATTATCAAAATCGGCATGGCGGCAGTACGTCCGGCAGAATTCATTATTTTACAGTTTACGCAGAATATCGCCCAGTAACTTAGGTCTATACCCTATAGATTTCAAGATGCATCGCGGCGGCAAGGGAGCGAATCCCCGGGAGCATATACCCAATAGATTTCAAGTTGCAGTGCGGCGGCAAGTGAACGCATCCCCAGGAGCATAGATAACTATGTGACTGGGGTAAGTGAACGCAGCCAACAAAGCAGCAGCTTGAAAGATGAAGGGTATAGATAACGATGTGACCGGGGTGAGTGAGTGCAGCCAACAAAGAGGCAACTTGAAAGATAACGGGTATATTTAATATGGGCGATTTATTGCCCATTTTTGTGAAAGGAAATGAGTTATGTCGCCAACGCTACCCGGTGTAACGATGACTCAGGCGCAGATAACAGCGTTCGGTGTCAGTACATTAAATATGCCCGTATTCATAGGGTATTGTACGAGATTGCCTGCCTTTTCAGCGCCTGTAAAAGTAAACAGTTTAGCTGAAACAGAACAAATAATAGGGAAAGAAGGGCGTTTGTATGCTCTATTGCGCCACTTTTTCGATAACGATGGGATACAAGCTTTTATTCTGTCGTTAGGCGCACCTGCTGGGGAAAATGCTAATAGTTGGCTTGAGGCATTACAACAGCCCGATTTGTATGCGGCTGTTGCAGCAGAGCCGCTAATTACACTTTTAGCCGTCGTTGAGGCAAGTGAACTGAACCAAAAAGAAGGTAATGAGGCTGTGGAAGCTTGGCGACAGTACTGGAAAGCAGTATTAGCGTTATGTCAGGCACGCAGTGACTTGTTTGCCATATTGGAGGCACCAGATGATACCGCATTAATCAAGCGTAGTTTGCAGGATTTTCATCATAAGGCACGTCAGTTTGGCGCTCTCTACTGGCCAAGGCTAGAAACATCTTATCAATCCTCTCAGTTAAAAATTTTGTCTCCTATTGGTGCAGTAGCAGCGGTTATTCAAAGTAATGATGTCCGGCGAGGGGTAGGACATGCACCTGCCAATATAGCGTTAAAACAGACGATTCGCCCGATAAAGTCCCGCCTGGAATTAGAAGAGTTGTATGAAGAATCGGATGGTTCACTGAATCTGATTTGTAGTTTTCCAGCTCGTGGTACTCGTATTTGGGGATGTCGTACGTTGGCGGGTATTGATTCACCTTGGCGTTATATTCAAACCCGATTATTGACTTCACACGTGGAAAGGCAACTCAGCCAGTTAGGGTGCATGTTGATGTTTGAACCTAATAACGCAGTCACTTGGATGAAGTTTAAAGGCCATGCTGGGAATCTATTAAGGCAGCTTTGGTTACAAGGGGTGCTGTATGGGCAGCGTGAAGATGAAGCCTTTTCCGTTGAAATAGATGAAAACGAAACGATGACTCGCCAGGATATTGATGAAGGCAGAATGATTGCTCGTATTCATTTGGCATTGTTAGCACCGGCAGAGTTTATCGCTGTGACTTTGAATTTTGATACTCGCTCAGGCATTGCGACGAGTACATAATAAATCGGAATATCTCCATGACACTACCAGCAGAGCTTTATACCCCAGCGGTTTCACATCGTTTTATTGTTAATTTTCTTTTTAAAGGTTTACTTCCTTCTCCCGTAGATATTCGATTTCAACGTGTTTCTGGTTTAGGGCGTGAGTTACAGGTTGAACAGCGCCATCAGGGGGGAGAAAACGCACGGAATCATTGGTTGGCTGAACGTATACAGCATAATAGCTTGATATTAGAAAGAGGGGTTATGGTCGTTACCCCTTTAACACTGATGTTTGATCAGGTGATGCGGGGGGAAACTCTCAATTGGGCAGATGTGGTAATTATTCTTCTCGATCAGGCTCAACGTCCGATAACAAGTTGGACCTTGAGTCATGCGCTACCGGTTCGCTGGCAAACAGGAGATTTAGATGCCAACAGTAACCAAGTGCTGATTAACACCTTAGAGCTGCGTTATGAAGATATGCGCATTATAGGGGTAAAATTATGACTATCGAAATCCGTGAACTCATTGTTCAAGCCCGTGTTGTCGGGACTGATACCAAAACAACACGAACCGTTCCTTTATCTATTGTGCAAATGGAAACACTTATAGAACAACGTCTGGTTGAAAAAGTGAAGCGGGAGATATTAGACGTACTCCGGGAAGAACAAGGTGGTGGGTTATGAGCTTGCTTGAACGAGGTCTGGCTAAACTCACGATTACGGGTTGGAAGGAGCGTGAGCGTAAACATCAGATTGGTAAACTAGAAGCAATGTATAACCCGGAAACACTTCAACTGGATTATCAAACTGATTATCTCCCTGATGTTAGCAATAATCAGGTAACAGTGAGTAACCGCTACGTTTTGTCAAAGCCCGCAGGGTTAACACTATCCTTGTTATTTGATGCCAATATGGCTGGTCTTACGACAACCGTCGAGTCCCAAATCACTACCCTCAAATCGCTTTGTTTAGTTAATGCAAGTACTGATGAACCCAATTTTTTGGAAATTAATTGGGGGGCAATGCGTTGGGAAAATAAAAATTATTTTGTTGGTCGGGCTAGTGGATTGTCTCTGACTTATTTGCGCTTTGATCGTAACGCAACACCATTGCGTGTGAGTGCGCAGCTCACATTAGTCGCAGATGAAAGCTTTGTGCTCCAGGATAACCAAGCCAAGTTAGATGCGCCGCCGGTATCAGTAGTTAATGTCCCGGATCTGACTTCATTACCTGCACTGGCGAATATCGCTAGCGTAACCACTATGTTGGGAGTGGATTATTTAATGTTAGCCCGCACCAATGATATGGATAATTTGGATGATATGCAGCCAGGTCAGACATTGCGAACACCGGAGGCATCATGAGTTTTTTAGATAACAGTAACTTCAAGCCATCAGATATCAAACTGTTCGTTAACATTCAGGGAGTGGAGAAGGAACTCAACGAACTGATAGTAAGCGAATTGAAAATCTCCCGACGTATCAATGCCATTCCGCAGGCAGTTGTAAAGCTAAGAGCGAAAGAGAGTGAAAGTGGTGTATATCAGTCTGATGTACAGCGGATGTTGAAGAGTTGCCGTCCGGGAGTAAAGGCAGAGCTTCGTATTTTGAATACCCGGCTATTCAGTGGCGATATTGTGCAGCAAAAAACAGAGTTAGTGTATGCGAAAACACACACTATCAAATTGGTGCTACGCCATGACTTACAGCGCATCACCGGTAATTTTCGTACCAGAGTGTTTGCGAATACCCGTGATCGTAAAGTGATAGCCGATCTATTGAATACCGCAACATTAAAGCCGGCATTTTCGGGGACATCACATTGGGATATAGATCATGAGCAACTGGTTCAGTATCGTTGCAGTGATTGGCAATTTTTGTTGCAACGGCTCTATGCTACGAATAGCTGGTTGTTAGCTGAAGAAGATAAAGATAACACTCAGGGGAAAGTGACCATTATTGCTCCAAATTCTTTGCCCCTGAATGAGCGTTGGACACTGCAACATCAGGCTGATCATCAGGCTATCCGGCTTTACAGCACGGAGCTGATGCTGGATAACCGGTTTGATACAGCGGAGGCTGTTGTTAGTGCTTGGGATATTGATGATCAGGCATTACTCGTGGCGTGGAAAGAAACCCTTAGTCAAGTTGGGAAAGATGCGTTAGCGTCAGATAATTTTAGCCAGACAAATAAAGATTCGAGTGAACTGTTATTAAGTTGTCCGCTCTCTACAAAAGAAGTTCAATTTTTAACGCGTAGCCAATTAGTCATGCGGCGCTTGACGGCCGTTCGTGGTTCACTGAAGGTTGAAGGCAGTACTAAGTACCGTTTAGGGCATGAACTGATGTTGTCAGGTTTTGGTGAAAATATGGATGGCTCACAAATACTGACGGGAGTGGATCATCGAATAACGGCAGAAGAAAGTTGGAAAACAACCTTACATGTGGGATTAGAACTGCCGTTAAAGGCAGAGTATGTCACTCAGGTTAACGGTGTTCATATCGGCAAGGTTGCTGATTATCAATCAGATAGCAAAAAATGGGATCGTATTCCTGTTTTGATCCCTGCATTTGGAACGAATATTCCCTTGTTTGCCCGATTGGGAAAACCCTACGCCAGCCACCAAAGTGGATTTTGTTTCTATCCTGAAACGGGTGATGAAGTCATTCTCAGTTTTTTGGAAGGGGACCCTCGTTATCCTGTCATTATTGATTCCCTGCATAATCCTAAACAACAGACTCCATTGCAAATCAGCAAAGAGAATAATCTCAAAATGTTGATGATTAAGCAGAGCGATAAAGATGAGCAACAATTGTTATTTGATAGCCAGCAACAAACAGTCGCGTTAATCGGTAAGAAAAATATCGAGGTTAAAGGTGAGTATATCAACCTGACTAAATCAAAGGGGACTCGATAATGGCAAATACGCTTATTGGCCAGGTATATGGTCAAGGATGGGCTTTTCCCATTAAATTTATTCCTGATAATAAAGAAACCGCAGATCAAACAGCCGGTATTGTTATGGCTCAAGGGATTGAAGATGTCAGTCAATCGCTGGAAATATTATTTCTTACCGAGCCTGGCGAACGAATTATGCGTGAAGATTTTGGTTGTGGTTTACAAGATTTTGTTTTTGAAAATATTAGTGATACGCTAATTTCTGCCATCAAAAATCGTATTCAGCAAGCAATATTACGTTATGAACCTCGCGCATATTTATTGAACGTTGATATTCAAACCAAAGAAAACCAACCTGGACATCTGCTCATTCAGATTAATTGGAAATTACGTGGTAGTGATATATCTCAGCGTTTAGACGGAGTGCTTAGACTCCATTCAGGTCAAGCATTGGAACTGTTATGACCAATTATATTATTATCGACGGGGATCTCATTCAAATAAATCCCAAATTTGAGGGTGATCGAACTCTTACGATTAATGGTATTCCTAAAATAAGCGGGAATGGAGATGCGCAAATTGAAGGAAAAAATATTTGTGTGTCAGGTGATCACTTAACTGTCTCAATTCCAGCCATTTATATAACCTCCAGACATCCTGTTGCAGGTAGTGGAAAAGTGAAAATTACAAATTTATCTGACGACCAACTAGCAGAATTTTGTGTTAGTGGGGATGTTGTGATTATTGAAGGCAGTCAGTTTGAAGCTCAGTTTACACCGGATAAGCCGGCCACTAATCCAAGTAACCAAGATGCAGATAATCCTGCGCCTTCGAATGGGAGTGGGAGATTTATACACTCACAGAACTTCGTTAAGGCAGAAAAATAAAAAATTTTGCCGAAGCGGTTAATAAGTATGAATAAGCGGGGCGGATAAAAACATGGATCTTGCTGAATTAAATAATACGTTGATGAATGACTTACCAACGACCAATTTTAAGTTAGAAACAAAGGACCCATTAACGCAATTAAAGTGGTTACAACGTTATACAGAAAATATTCGTTTTTATGCGAATGATGATTATTTCTGGCATCAATTCTGGTTCTTAAAAAATCACACACCAGAAGCGCTCTTTGCTCGTTTGCAAGGTGAAACGTTGGCTGATGGAGAATTGCCTCCTCATCAAGCGCTATTGCTGGCCTTTTTACAACAGCTTAAGACGCCAGGAATCATGCTTGATACTTTTTCAGCCCGTCATCGGCAATTGTACTATCAGGAATTGCTAGGGATAACGCAGAAAGATGCACAACCTGATCATGTGGCGCTTGGCGTGGTATTAAGTACTGGTATTGCAGAATATTTATTACCGACAGGCACATTAGTGGATGGTGGACAAGACAGCAGCGGAAATTCACTGCAATATGCGTTGGATACCGATTTATTGGTTAATCCAGGGCAATTAACAGATGTTCGCTACAGCTATTTGGATCATAAGACCTATAAAATCTTCATCTTGCAAGATGATAAAGCGAATATCAGTTGGCCCTCTTCAGGCGCTCGTTTATTTGTAGCACCTGAGGGCAACGGACAGGAAAAGGCACCTGAACAAAAGTTGGCACTTTACCTGGGATTTGATGATATACAGCCAGGGCAAACTCTTTCTTTATTTTGGCAATTCATTGCATCAACTCCCCTGACATTAAAATGGTTTTATCTGAACGAGATAAATAACTGGGTGAAGCTAGATAGTGTCAGAGATAACACGGATGGCTTTTTTATCAGTGGATTATGGCAAGCGATATTACCTGATGATGCGGTGAAAATGTATTTTCCAGAGACAACTTCTGTAAAACGCTACTGGATTAAAGCTGAGGTGGAATCGCTTACTGAATCTGGCGATTTGTGGCAACCGCTATTAGAAGGCATCTTGTATAACGCTCAAACAGCAACGCTGGTTGATGCAGACAACACAGATGAAAAGCACTTTCATGATGGGCTGATGCCTTTTAGCGTGCAGCATTTGGTCAACACCGTTTCAGAGGTAAAAAAAATTGAGCAGCCCTGGTCTTCTTGGGGGGGAACGCCACAGGAAGACACTACTGATTTCTTCCATCGAGCGGCAACACGTCTTCAGCATCGCCAGCGTGCGTTAACTTGGGATAACCAAATTGCCATGTTGAAGGCTGAATTTCCGCGGATTTATGATGTCATCTCACCAAATATCACGTGGATGAACCAACTTCAGACATCAAATACGCAAACGCTGATCGTTATTCCTGATGTGAACTACAGCGACAACAAGGATCGCTTACGGCCACAATTCAGCCCTGCCAGCTTGCGACAAATGAGTGACTGGTTACAGATTCACACTAGCGCATGGGCGAATCCACAAGTGGAAAATCCAATTTATATTGATGTCTCTGTGACCTATGAGGTGCAATTTAGTGCGGGTGTGAATCCTGATTATGCCCTCCGGCAATTACAACAATGGTTGAGTTCAATTTATATGCCATGGTATCACGCAGATAAAAAAGGTGTTGCCGCTGGCGATCAAATCGATTTTTACCAACTGTTTGCAGATATTCAGCGAGTACCTTACGTGGAGCATGTCAAAACATTGACATTGACCACAAAAGACACCTCATTAACCAATGGCGGGGTTATTAAGGCACAGCAAAATGAAGTGCTGGTGTTGGTATGGCAACAAGGAGAACAAATTAGGCAGGGAGAATCGAAATGAGGCAGCATAATGAGTTATTTCCTGTAGTAAAAGACGCGATAAGCTTTGAAAACCTGCAAGCTCAGGGTGAGAAGGTTATTAGTGATCAGTCCGGTAACATATGGAGCGATAAAGATAAACATGATCCTGGTATAACATTACTAGACTCTTTAAGTTACGGTGTTTCGGATTTAGCGTATCGGCACTCATTACCTTTAACCGATTTATTAACCATTGCTGGAAAAGATACGCTTTTTCCAGCCGAATTCGGGCCACAGCAGACGCTAACTTGTGGCCCTATAACACTGGATGATTACCGGCGTGCGTTACTTGATTTACATGGTAATGATGCATTTAAAATATCAGCTAGTGACCCCAGAGACTTTTTGTTTCAGGATATACAGTTAATTTGTGAGCCAAAAAGTAAGCGTTATAAATACTATTTCAATCCCGAAACGCTTGAATATACATTCACGCCACCTTCAGGGGATAAATTTAAAACTTTAACACTACGAGGGAATTATTGGCTTTATTGGATACCAACCCGTTGGGCAGGTAAATCAGCTAATTTGCCGTTAGTTAAGCGGGTGATGGAAGATTTTCTCCGTGAAAATCGAAATTTGGGGGAAAATGTTGTTCAAGTGACACGGGTGATATCAACGCCTATTTATCCTGAGCTGGTCATTGAGCTGGCGGATGATATTACAGATGCGGCATCAGTATTAGCATCAATCTATATGCTATTAGAACAGTGGGCGATGCCGATGCCTGCTCGCTTTACTACCGAAGCATTACAGGCCAAGGGATTAACAAACGAAGAGATCTTTGATGGGCCGTGGTTGCGTCATGGTTGGATACCTCAGTTACCGACCTCTCAAAACTACCATACAGGCATGGTTCTGAAGATGAATCATCTGATTAACCAATTGCTGGCGGTTGAAGGTATAAAGCGCGTAGTTAGCCTGACGTTGCCAGAAACAGAATATTTGCATCAGATAAAAGATGATAATTGGTCCTGGCAATTAGATGTTGGTTATTATCCATTATTATGGGGAGCTAATCCACTAGAGGTAATTACAGAGAAAAATAACAATTATGTCAAATTGTTCGCAAAAGGTGGGGTACGATTACAACCTGATCAGAAAAGTGTTGAGCGGTTATTATCACAGGAATCACTCATTAATAATGCTGCATCCACGTTACCGGCTGGTAAGGTGCGTGATCTCAAAGCCTATACACCTATAAGCCGCAGGTTGCCTGCCTGTTATGGTTTGCAGAATACTTTGCAAAAGTTAAAACCTGAACAACGACACTTATATCAGTTCCTATTACCATTGGAGCAAATGCTTGCTGATGGATGTGCGCGGCTTGCATTTTTGCCACATTTGTTAGCATTTAGGGACCGAAGCGGAAATATCAGTGATACACTCTGGCCTTTCAAGAATACAGAGGACACAATTGCCCAACAGGTTCATCAGGAATATGCCGGTACATTAAAAGCCTTTCAACAGCAGGAAATTAGCCTGTTTGATGATAAAAATAGACCGCATCATGGCAATATCAATCGGGAATTAGATATTCTTGATTATCTGCTAGGGTATTTTGGTACACAACGTGCAAAGCGTCCATTAACGCAGGATATTCATGATTTTCTGCAAACCCAGCGAGGTTATTTGGCACAGCAGCCGGAGTTGGGTTATCAGCGTGATAATATCCGTATTGATCGAGTTTCAGCTTTACAAAAACGTATAGCAGCCCGAATTGGGCTAGATGGTACTATTTTCAAAGAATCGGTTGATTTAAGTAAGTTACCTTTTTATTTGATTGAACATCGTCAGCTTTTACCAAATTTACCCCATCTTGACTTTCAACATGATCAAACTCCCCAATCTTTTGTGATTTCCGACAACATTGTTAAAGTGAAACAAGCGGGAATAGCAGATAAAATCGTTCGTGGACAGCTTATTGATTTTATAGATATTGAAAGCAAATTTACCGTTCGTGCCCAAATGATTGTCGCTGTAGAGGGAAATGAATTTTCTCTGGATACAAAAAATAGTATTCAACTTGAAAAGAATCTGCAGTTATTACAATCAGCGTCTGAGAAAAACAATTTACGATGGAGAAATAGCACGGCGTGGTTAGAGGATATGACGTATCGTATCAATTATACTGACGATCAGGTTATAGACGATAAAACAAAACAATGTCGTTTACAAAGTAATACTAAATCGCCTTTTCCAGCCTTAATTGCACCAAAAAATAAGATTACGATTATTAAGCAATCTTCTCCACTCTCCAGTATTGCTGAATTTACTGATGAACCAGAATTCAAATTAGTTGCAACGGTGACAGAGATTGATCGGATTGAAGGGATATTGACTATCGAACGGGATGACAACCAACTCCCTTTCCCGACTAAAGAAGAGAGTAATCAATATATATGGTACATATCTGATGAAAACTATATTTCAAGTGATCGTTTCTCTTTTGTGGTGAGCGTCGTGCTGAATCGCGGTTTGGTTGAAAGGGAAGATATTGATCAATATAAGCTAGAGGAATGGATAGAGCGTGAAACACTTGCAGAGTTTCCTGCACATATTTCGTTAATTACTCATTGGCTGGCATCTGAAAATTTCGATGATTTTGCGAAGACATATCAACGTTGGCAAAACAATGGGGCGCAGTTAGGGGATGAATCCTACACCATTTTGGAAAAACTGACATTAGGGCATTTACCAACAGGACTTACTGGCATTAGTAATATGTTTATTGCTACAGAAGCTCAGCGTCTAGAAGTTGTTGGCGAGAGTGGTAATGAGTGGAATACCCAGGCAATTATTAACAACGAACTATTCTATGTTCCCTCACAGAATAGTTAATACCGAGTGTTGTGATCAACTTTTATTATAAGCCGGAGGATAAATGGACAACAAAAATAACAAACCTACTGATCAAGAGATTCTAAAAACATCACGGGCTGTCGGAGAAATTCCTTCAGCGGATAATTTAAAAAATCGTTTTAAAGCTCGTTCGATTCCATTAGAGACGGATTTTACTAATCTCATTGACCTTGCTGAAGTTGGACGATTGGCTATCGGCCAGTCACCATCGCAGCAAAGTAAAACGCCTGGCACCGGAATGGAATTAACTTCGGATGGTAAATTACAAGTCAAGGCTGGGGCAGGTGTTGATATCGATAATAATAATCGTATTACTATTAAGTCTGGTCATGGAATTAAGGTTGATGGAAACGGCATTTCCGTTAAACCAGGTTCGGGTATTAAGGTTGATAGTAATGGTGTAAATGTCAATATTGATGATTTTTGGGAGGAAATACGCAATAAAATTATGCCTAAAGGAACCATGCTGCCTATTTATGGCACACCTAACCCCTCTGCGCTGCCAACAGGATGGGAATGGTGTGATGGTAAAGATGGCAGACCTAATTTAAAAAAAGGGAAATATAACTTACTATCAGGTCAGTCTTCAGGTACTGATACTTTTTGGGCAGATAATAAGAATGGAGATACAGAGATCAACGTGTTATTTGTTTACTATATGATTAAGGTTGTGTAATATCTTAAGTAATATGCATTACTCTAAAATGAATGATTTATATTTAAGTAACATAATAATTAAGTTGTGTTGTAGGGCTGTTTTTATGAGAAATATAAAAACGGAGGTAATAATTGGCTTCAAAATATCAGTGATGAAATAGAGTTATTTCGCTTTATAAAAATTTTGTTTTATTTCTTTTAATAATTATTTATAGAAGGTAATGATATGTGCACACAAAAAAACGTGTTAGATAGACTGAAAGATAGAAATATTACATTGGGTTGGGATGTTGTTGTTGCATATAACCAAGAAAGTGTTAATAAGTTATTGAAGCAACAATATGTTGAAAAAGTTTACTCAAATGAACATTTTGTTTTTAAAGATTGGCATGATGATAATAAAACGAAATTTATTGAGGGATTAACAGTAGGCGCTCCACTAGTTTCATTTGAGGAGGCGTCTTTATCCGATGCTAATGTAAAAGTGACACTTAACTTTCTTTCTGGTAGATGGAGAGTTATACAAGCAAATACCGGCACACCAATTGAATGGAAAGAAATTGTTCCTGGCAGTGGCTATAAAGCAGAATTAGTTGTTCCGCTTAAATCAATAACTGGTAGTGTAAGTAAAAAAGATATCATATTAAAATTCAAAGATGCTGTCGTAAAAAAAATAAATTTATTTGACAATCAAGAGCCTGATTTTATTAATTATTTCAAGCAATCGATCAGTGAGGGAAATTATACTTTAGGGCAACTGGTGACAGACAGCACACCGGGATTAATTCCTGCTGAATTTCATATTCGTACTCAACCCCATCCAAAAACACGTGAGCGTGGTTCTCAATATGTAGGAAATGGTGCGGTACTGTTGTTTATTAAAACGCAATATGGCGGAAGTGGAACATTGCCTGTAAATGATTTTGATTGGTTAATTCCTGATGATCATACTAGCGCATTAGTCATTTCGAGTAAGACCATGATGGGGCAAATATTGCCAAAACAATACAAAGATAAATTGCCTGGTGATCCTCAGTTTAGCCCACCAAAAAGAGTCAATGATAAACAAGACTCTGCTTATTATATTACGATTACCGATGGTGGATTTGATGGTAATAGCCCTATAGAGAAGTCATGGTTACGTTCTGATTATAGCAATGGGATTTGGACTGGTGAACGTGGTAATGCTATTATTGGTGAAAAAGGAAAGCGGATACCACCACGTTTTCCATACCAAAATTTTGTTATTAAACCTCATGGTGAATCGTTATTTCAAGGATGGGAGAATAAGATAAATTACACTCAAAAGTGTGCAAGATATTTCCGACATCATAGTAATAGTATAACTTTCGAAGATACTGCATTAATGGATCTCAGTATTGGTGGACAAGGTAGTATCAATTGCCAGATTGATGGTGAACATTTCTATTTAAAATCAGATGATTTTTCCCCCAATGTCAGCTATGAACCAACTTCATTCTGGGATAAATTTATCGGTGGGGTGGATGCAAATGTGAAAGATGAATTCAGAGATGAATTAGCACAACAGGCAGAAGCAAAGTTAAAACAGGTATTTAATATTGAATTGCCTGAAATCAGTCTGTTTTCTATTAAACATCTGCTCTTTCCTGGCATGGATGTTATGCAACTTAAACAGGGTTATTTCCCAGGAGATTTGATTATCTTTGGGGATATTTCACCTAAATTGACCACAATTCAGGTGGCTCCTTTGGAAGCCATGGTTGCCCTTAAAGAAAATCAAAAATTCACTGTCGTACCTGAAAATAAAAATGTTAGTTGGAAGTTGGATCATAATAGTGAGGCTATCAATGATCCGGGAAATATTGATGATAAAGGTATTTATACGGCACCGGGCAGAATCAGATCTGGTTCTGAAGTCATTAAAGTCACTGCAACTGACGGCGATGGAAATCAGGCATCGGCGGCGCTGACGTTGGTTCCTTCTTCTGTTGCATTAACACCTTCTTTTGCTTTTATCTCTGAAGCAGATAAGAAACCTATATTATTATTGGCGAATGTCCTAGACGGAAAAGCAGTAACATGGAATGTGGAAAGCTGTACAGGCAGCCAATGTGGTTCTGTTGATCAGAATGGGCTTTATACTCCACCAGCAGGGCGTTTTAACGATGGATTTACTTTTGCATCCATCACCGCAACTGCAAAAGATGGTAGTCAAGCACGAACCATTATTTGTCTAATGGCATCAATGCCAGGACATGGTTTTTACAAGGTTGAACCTAATTTACGTTTGAATGTGAAAGTAGGGGAAGAAATTATCTTTAAAGCGCAGGCAGATAGCTATAATGGTGATCCTGATACTTGGGAAATTTTCCCTCCTCGCGGAAAATTAAGTGAACCTGAGTTTGAACCCAATAATGATCCTGAAACTAATGATACAATTTTTGGTCATTATAAGGTGACCTATACCGCGCCGACTAATGTTACCTCACCTGAATTGCTTGTTGTCCATGTATGGGAGAAAAATAGGCATAATGAGAAAAACAAAGGTAAGGCAGGATATGCACTTATTGAAATTATCCCAGATGATAAATAGAAAATTTATTTAAATAAAAATCACAGCGGGTTTATCTCGCTGTGATTAAAGTCATCTTTTTTTATAGATTGTTTATCTCTAATAATAATTTTATTTTATAATATAAAGGAAATTAAAATGAATAATGAATATAAAAATAACACCGTGAATTGGCGTATTTCACCTGATACGGTAGGAAGTATTGATAATAACGGTTTATATACAGCACCTAATCGGGTAAAGAATATCGAATTTGTCCAAGTAATGGCAAGCGATGCTAATAATAATCAATCTTCTGCGATTATTACTGTTATTCCCTCTTCTGTTGCGTTAACGCCATCGTTTACTTTTATCTCTGAGGCAAAAAAAACATCAGTCACTTTTAAAGCGACAGAACTTGAAGGGAAAAAAGTGACATGGAGTATAAATAATTATACCAGTAATCAGTATGGTTCCATCGATCAAAATGGTATCTACACACCACCGGAAAGTCGTTTTAACGATGGATATACTTTTGTATCTATTACAGCAAAAGCGGAAAATGGCGCTGAAGCGCAAGCGCTTATTTGCTTGATGGCCAAAATTCCAGGGCATGCCTTTTTCGATGTTCAGCCTAATATATGTTTAAGTGTGAAGCCTGGAGAAGAAATCATTTTTAGAGCTAACGCAGATCGTTATAATGGTGATCCTGATTCCTGGGAAATTTTCCCGTCTCTTGGTAAATTGGGTGAGCCTGAGTATATAAAAAATAACGATCCAGAAATTCCTATTTATGGATATTATCAAGTGAAATATATTGCGCCAACCAATATAAATTCTTCCCAAATACTCGTTGTGCGTACTTGGGAATATGACAAACATGATGAGCATAATCAAGGTAAAGCAGGATATGCATTCATTGAAATTGTGCCAGAAAATGAGCTTTAATATATATACCCAATAGATTTCGAGCCGCAGCGCGGCGGCAAGTGAATGAATCCCCAGGAGCATAGATAACGATGTGACTGGGGTGAGTGAACGCAGCCAACAAAGAGGTAACTTGAAAGATAATGAGTATAAATGACTTTAGTAAGAGAAATTATGGCTTCATTCAGAACTATTTATTAGAGTAATTAACTTTATAAAGACATTTAATGGAAAATATAATAGAAAAATTTAATATTAATATTGAAGTCTCATCTGAAATTATTGGAGAGAGTTTATTAAACTCCCCTTTATTGATGAGTAGAGAAATCAGCAATCAATTATCTGAAATATTATTAGATTATAAAGAATATAATATTGCATTGGATAAGTTAGTGTTAAATATAGGAGAAATACCCTATGAAATATTTGAACAACAATTCTATGGTCGTTTGGGAAAATTATTAAATGAAAAGTTAACAATAATAATAAATGATAAATTATTGGTAAAAAACATATCAACCTCGTTATTTCCTGAATGTTTTAGTGAAAAAAGAAACCCATTATTAAATAGAGTCATAAAAAATTTACCTTCTAATTTGGTTTTTGAAGTTCATTCAATGGTAAAAATAGAATCAGTAAATAACAAAAAACAAGCTAATATATTGACATCTTATCTGGCTTATTCTTTTTTTAATAAAAGCAAATTACAACAACATTTATTTTCCACTAGTAATAATAAATTAATTGAGAGCTTATACGCACTTTTTCTAACGGATCAGAATCGAATACCTACTGCTCATAAAATAGGAAAAGGTGCACTTATACTATCTGCCCTTATTTGGCTTTATTCTAATTCCAATGATTATCTGCCCAAACCAGAAAGCACTCTGTTGTTACAAATAGAACAGGATATAAAACAAGGATATTTGCCTTTAACGTTGTTAATCACTTTCTTCCAGAACAGAAATGGCGGGCGTGTTTTTTGCGATTGGCAGTATGCGTTATGGCAAATCGATATCATCAAAAATCACTTAGGCATTAAAATAACATCGAAAGAACCCCATTTACGGGAGAAAATAATGTTACAACCAGTTAATGCTTCTGATCGATCCTCTGTGCTGATATCAGACGAAAAATTGACAATACCGTTAACAATTACAGGTGCGGGATTAGTGCTTCTCTGGCCACTATTAACTCCACTATTTTCGTCTTTTGATTTGTTAGATAAGAAAAGTTTTTCAGACAATTTGGCACAGGAAATAGCATTTAATTTATTGGAATGGTTAGTCTGGGGAGATGAGATGCTGTTACATCAGGAATCATCATTATCTTTATTACTCTGCGGAATAGATCACCAAACAATACTGGAGCGCCAGGTTCTTATTCCTGAGCACAAGGAAAAATTAAATAACTGGTTGCAAGGTATTTGTACTCAACTTTTCTCTTGGAAAAAGCTAGGGATCGATGATATGCGCCAACTTTTTTTGCAGCGTCAGGCTGCACTTTATTATGAAGATGATGGCCGTTGGTTATTAACGGTGCAGCGTGAAGCTTATGATGTATTACTGACTCAAATGCCTTGGCCGTGGCCATTGAATATTGTGACATTACCTTGGCTAGCTGAGCCGATTAGTATCACTTGGGAAGGTATCTCTGAACCAACGGATTTGTCATTTTGGTAATCCAATATCTCATTAGGAACTCTATGCATGTACGATTTATCTGATGATCTTGCCAGACAGAATATTTCACCGGAATATGAATTGACGGTTTTGCTGTCTCAGACTGCTATATTGGATAAACGAATTCGTTTACGAATTCAGGAATTAATGCAACAGCAAACACTATTGGGAGAAAGTGGACAGACGTCTTTTGATGATATTTCATTTTCATTCGTTTCGAGTGAACAACAAAAATCATCTTATTTGGTGTCACCGCATCAAAATTGGACGAAAGAGGATTTTCCTCCTGAGCCGATCCCATCTCGTAGCCGTCTAGGACAATTAGTTGAACGGTTTGACTTAACTCAATTTGAAATTGATTTGATTTTATTGTGCCTGTTGCCTCATCTTGACAGACGTTATCTAACGTTATTTTCTCTTGTTCCGGTAAGTGGAGGTAATAACAGCAAAAAGCAGATGTTAACGTTGGGATTGGCTTTGGAGTTGCTTTGTCCGAGTGTAGTAGAGCGCAATGCGCAACGTGCCAGTTTATTACCACAGGCACCGCTTTGGGATTATCGTTTATTTCAGTTGCGCGGTGATATGTCTGTTTCCTACGATGAAATACCGTTAGCAATCGATAATTCTCTTATGCATTGGTTATTGGGGCATGATGCTCTCCCGATTTCTCTTCTCTCCCGGGCTCATTGGCTTCCTGTTCCTGAAGTGCCTGATATTTTGCCTGATTTCACCAACCAATTGATAGAACTCTGCCAAATGGAACAAGAGGGGATGCTGACAATAATCGCCGGCGGAGCCGGAAGTGGCAGCAAAACAAGTGTTGCACGCGCAGCATCACAAGTAGGGCGCTCTGTATTGTTGTTATCGTTAGCATCAGTGACACTGAGTGAACATGAAACTATTACACTGATAACACTGGCATTACGTGAAGCACAACTAAGAAATGCCTGTCTTATGTTTGAAGCTTTGGATGAGTTTTGTGAAGCACGCCCCGCTTTGCAGCTCTGGCTAGGAAATCGACTGGCTCGTTGTTCGATTCCGCTGTTTTGTCAATTACCTAAGCAAGCATCATTATTGCCATTGGATGCAATTTCACAAGTTGTATTGTCTATGCCAATGCCTTCTTTAATGGTGAAGGCTGCAGCATTAGCTTCAATGATGACGAATTATTTTCCAGACAATTCATTGGATGTTGAAAGTTTAGTGACATGTTTCCATCCTTCTCCATTGATATTGAAAAAGGCCCTTAGTGAAGCAGAAATTTATCGCCGACTACGGGGGGAAACGGCTAGTTTGAGATTAGATGATGTGCAAATGTCCCTGCGTTTTCGGTTACAGCAGAATTTTGGACGTTTAGCACAGAGAATTACACCACAACGAACCTTTGATGATTTGATCATCAGTGAATCTCAACAGCAACAATTACAAGAAATCCTGGCGGCTATTCGGCAACGAGATAGGATGCTAGAGCAAGGATTTGCTCGTAAAGTGAGCTATGGGACGGGTATCAGCACGCTATTTTTTGGTGAATCTGGCACAGGAAAAACGATGGTAGCAGAAGTGTTAGCTGGTGTTTTAGGTGTGGATTTGATCAAGGTAGATTTGTCCACTGTGGTTAACAAATATATTGGTGAAACTGAAAAAAATCTGGCTCGTGTTTTTGATTATGCCCAAGAAGACGCCGGGGTATTGTTCTTTGATGAGGCAGATGCATTGTTTGGCAAACGAAGTGAAACTAAAGATGCAAAAGATCGTCATGCTAATATTGAAGTTTCCTACCTATTGCAACGCCTTGAAAGTTATCCAGGGCTGGTGATATTAGCCACCAATTACCGTAATCATTTAGACTCAGCATTTAGTCGTCGCCTGACTTTTTCGGTACGATTCTCTTTTCCAGATGTTTCCTTACGGGAACGGATGTGGCGGATTATCTGGCCATCGGGAATTCAATTAGCCGACGACATCAGTTTTTCAGCGTTGGCAAAACGGGCTGAATTAACGGGGGCGAATATCCGTAATATTGCGCTACTCGCTAGTTGGCTGGCAGTAGATGAAGGAAATGAAAAAATTACTATGGCTCATATTGAATGCGCATTACGACGTGAACTGAGTAAAGTTGGGCGCATTGATTTACCTTAATTTTTCTTTGTAATCGGGAGACAACTATGGTTAAAAATATCAAATCAGATGAAACCTTACTGATATTAAATAGTAAAATAGAAGATGCATTAAAAGCGTATTTACCGGGCGAAGATGTCGTTATTCGGTTCGATATGTTTGGTAAAAATGAAAATCCAGATTCTCCTACCGTGTGCGTTTTTCTTTATGATATTCAGGAAGATCTGCAATTACGCGTGGGAGAAGGGCGGCAATACCTGCCTGCGACAGGAAATTTTGTCCCGGGATGTGTCAATGTTCGTTGTAATTATCTTATTTCCTACTGGGAGCCGGAACAGAGCGGAGGGCAGGGATCGCCAACCATACGTTCTAATAGTCAATCAATGAAGATAATGAACTGTGTATTGAATGCATTAATTAATCATCGTTCATTTCCTGGTTTACCCAGAACTTATACGAGAGTTCTTCCTCCTAATGAACAATTAAATAGCTTAGGAAACTTTTGGCAATCATTAGATAATAAGCCTCGACTATGTTTAAGTTATATGGTGACTATTCCTATTCAACTTACCCCGCCGACAGAGAAGGTATCTCCTGTCATTACCTCAAAAACTGATATTACTCGAAAACCATCGCTTAACTTTTATCTTGAGGCAGATGAAATTATCCGTCAGGCATTAGTTGATGCCTTAATATCTCAAACAACAGAATCTATGGATACGATAACTAGCTGGCTGGCAAAAGTTGTTATTATTTGTCGACCACCAGAAATAATGAATAAACAAATGATTGAACAAACTGTGAAATTAATTATTGCTGGAATTACAGAAGAGGGATTAGCTGGAAATATAAAGACAATCACTCAAAAGTGGGTGGAAGAGAAGACGATTATTGGTGAAATCGACGATGTTTCTCTAGTTATTTCCCAAGTTGACACGACAGCGTTGTCTGCTGTAACAATACCGACATCTGTTTAA
SEQ
ID
NO:96(Pnf表位)
TGQKPGNNEWKTGR
SEQ ID NO:97(PVCpromF)
TATCATATGTCTACAACTCCAGAACAAATTGCTG
SEQ ID NO:98(PVCpromR)
ATCTCTAGAACAGATATTCCAGCCAGC
SEQ
ID NO:99(ParaINF)
GGCGTCACACTTTGCTATG
SEQ
ID
NO:100(ParaINF)
TCGGTGGCAGTAAATTGTCC
SEQ
ID
NO:101(F1引物)
ATGTCTACAAGTACATCTCAAATTGCG
SEQ
ID NO:102(F2引物)
GACTCCCTTGAGGGTACGG
SEQ
ID
NO:103(F3引物)
TTCTGATGAGAGTGATGGTAC
SEQ ID NO:104(F4引物)
TGAATAAAGAATTCAGTCAATATC
SEQ ID NO:105(R1引物)
TAGTGGCTGATGAAAGTCTG
SEQ ID NO:106(R2引物)
GGAAGCCAAAGATAATGAAGTG
SEQ
ID NO:107(R3引物)
CATTTCTTCCCTATGGTTG
SEQ
ID
NO:108(R4引物)
TTAAATTCCTACAAGATTATCTTT
SEQ ID NO:109(tBid氨基酸序列)
RSSHSRLGRIEADSESQEDIIRNIARHLAQVGDSMDRSIPPGLVNGLALQLRNTSRSEEDRNRDLATALEQLLQAYPRDMEKEKTMLVLALLLAKKVASHTPSLLRDVFHTTVNFINQNLRTYVRSLARNGMD
SEQ
ID
NO:110(E.coli序列优化的tBid碱基)
CGGTCAAGTCACTCGCGTCTGGGGAGAATCGAGGCTGATAGTGAGAGCCAAGAGGATATCATAAGAAACATAGCACGCCATTTGGCACAGGTAGGCGATTCTATGGATCGCTCCATCCCGCCTGGACTTGTCAATGGTCTTGCGCTTCAACTTCGTAACACTTCCCGGTCCGAGGAAGACAGAAATCGGGACCTTGCGACTGCTCTGGAACAACTGCTTCAAGCATATCCTCGTGACATGGAGAAAGAAAAGACTATGTTAGTATTAGCTCTTCTTTTAGCTAAAAAGGTAGCTTCGCACACTCCAAGTTTATTGCGGGACGTTTTTCACACCACTGTTAATTTCATCAATCAGAACCTGCGTACTTATGTGAGATCTTTGGCGAGAAATGGTATGGAT
SEQ
ID
NO:111(BaxBH3肽(aa59–73))
LSESLKRIGDELDSN
SEQ
ID
NO:112(E.coli序列优化的BaxBH3碱基)
CTGTCGGAGAGTTTGAAGCGTATAGGTGACGAGCTGGACAGCAAT
序列表
<110> 华威大学（THE UNIVERSITY OF WARWICK）
<120> 前导序列
<130> P61200WO
<150> GB1908155.3
<151> 2019-06-07
<160> 112
<170> SIPOSequenceListing 1.0
<210> 1
<211> 299
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 1
Met Met Arg Glu Tyr Ser Asn Glu Asp Asp Phe Ile Lys Glu Lys Thr
1 5 10 15
Asn Leu Val Lys Ser Glu Asn Val Glu Ala Asp Asn Tyr Leu Glu Thr
20 25 30
Glu Tyr Leu Thr Tyr Leu Ala Lys Leu Ile Gly Met Thr Glu Arg Glu
35 40 45
Asn His His Leu Asn Ser Ile Lys Leu Ile Asp Asp Ile Ile Glu Leu
50 55 60
His Asn Asp Arg Lys Gly Asn Lys Leu Leu Trp Asn Asp Asn Trp Gln
65 70 75 80
Asp Lys Ile Ile Asp Arg Asp Leu Gln Ser Ile Phe Lys Lys Ile Asp
85 90 95
Glu Met Val Ser Glu Phe Gly Gly Leu Glu Ala Tyr Lys Asp Ile Val
100 105 110
Gly Glu Asn Pro Tyr Asp Pro Thr Glu Pro Val Cys Gly Tyr Ser Ala
115 120 125
Gln Asn Ile Phe Lys Leu Met Thr Glu Gly Glu Tyr Ala Val Asp Pro
130 135 140
Val Lys Met Ala Lys Thr Gly Lys Ile Asn Gly Asn Gln Phe Ala Glu
145 150 155 160
Lys Leu Glu His Leu Asn Ser Ser Asn Asn Tyr Val Ala Leu Ile Asn
165 170 175
Asp His Arg Leu Gly His Met Phe Leu Val Asp Ile Pro Ser Thr Asn
180 185 190
Arg Glu Arg Val Gly Tyr Ile Tyr Gln Ser Asp Leu Gly Asp Gly Ala
195 200 205
Leu Pro Ala Leu Lys Ile Ala Asp Trp Leu Lys Ser Arg Gly Lys Glu
210 215 220
Ser Ile Asn Val Asn Lys Leu Lys Lys Phe Leu Asn Asp Glu Phe Thr
225 230 235 240
Met Leu Pro Asp Asn Glu Gln Lys Gly Leu Ile Ala Glu Ile Phe Asp
245 250 255
Leu Asn Lys Asp Ile Asp Ser Val Lys Ser Gly Lys Ile Lys Lys Asp
260 265 270
Lys Ala Val Asp Ile Tyr Leu Arg Glu Tyr Asp Ile Asn Asp Phe Ile
275 280 285
Ser Asn Ile Glu Lys Leu Lys Thr Lys Leu Ala
290 295
<210> 2
<211> 309
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 2
Met Phe Gln Asn Arg Ile Arg Asn Glu Lys Thr Thr Gln Ser Gly Lys
1 5 10 15
Gly Lys Thr Leu Asp Arg Met Thr Asp Ser Leu Tyr Leu Glu Ile Pro
20 25 30
Asn Val Glu Ala Val Thr Leu Ala Tyr Gln Lys Leu Thr Ser Lys Tyr
35 40 45
Arg Lys Phe Asp Asn Lys Thr Lys Leu Ile Leu Asp Ser Ser Asp Glu
50 55 60
Phe Ser Gln Leu Lys Ser Glu Lys Gln Arg Lys Gly Phe Ser Lys Ser
65 70 75 80
Gly Leu Lys Asn Asn Gly Val Ser Asp Arg Lys Phe Ile Tyr Thr Lys
85 90 95
Asn Ala Leu Lys Asn Phe Ala Ala His Ala Gly Tyr Glu His Asn Gly
100 105 110
His Tyr Glu Asp Glu Phe Val Asn Phe Lys Asp Asn Asn Lys Asn Leu
115 120 125
Ala Lys Gly Lys Leu Phe Pro Gly Ile Ser Leu Ile Glu Arg Arg Lys
130 135 140
Leu Ser Ile Val Lys Asn Lys Glu Gly Lys Trp Glu His Lys Glu Thr
145 150 155 160
Asp Glu Ala Glu Ala Tyr Lys Val Thr Asp Ile Glu Lys Phe Ile Ser
165 170 175
Gly Val Arg Ser Met Tyr Leu Gln Gly Asn Thr Phe Leu His Ala Lys
180 185 190
Thr Glu Ala Leu Ile Arg Lys His Ile Ala Asn Asn Glu Asn Ile Leu
195 200 205
Pro Thr Met Ala Gly Ile Ala Gly Leu His Ala Glu Val Gln Ala Leu
210 215 220
Asn Asn Leu Phe Ile Ser Gly Asp Lys Gly Thr Lys Lys Arg Glu Lys
225 230 235 240
Trp Lys Tyr Ile Arg Asn Met Leu Glu Ser Ser Ile Phe Thr Gln Arg
245 250 255
Leu Thr Thr Gly Gln Ala Gly Lys Asp Phe Ala Ala Cys His Asn Cys
260 265 270
Ser Gly Ile Leu Ser Ser Pro Val Asn Val Ile Thr Gly Lys Val Glu
275 280 285
Ser Ala Gly Asp Asn Phe Leu Ser Thr Leu Ser Arg Tyr Lys Thr Ser
290 295 300
Gln Glu Ser Pro Ile
305
<210> 3
<211> 278
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 3
Met Glu Arg Glu Tyr Ser Glu Lys Gln Lys Asn Pro Ser Lys Leu Ser
1 5 10 15
Arg Lys Thr Ala Ile Ser Glu Arg Ile Ala Ala Leu Glu Arg Ser Gly
20 25 30
Leu Ser Asn Ser Asn Gln Pro Val Pro Gln Phe Ala Arg Pro Tyr Thr
35 40 45
Ser Asn Arg Pro Val Val Asn Ile Asn Pro Gly Arg Ser Ser Ile Ala
50 55 60
Val Ala Thr Ala Asn Ser Thr Ser Pro Val Asn Ile Pro Thr Pro Ala
65 70 75 80
Pro Ala Ser Pro Asp Lys Leu Leu Pro Ser Thr Ser Cys Asp Thr Thr
85 90 95
Ser Ser Ile Leu Ile Val Gly Lys Tyr Asn Leu Glu Leu Thr Ser Gln
100 105 110
Gly Lys Ile Val Val Phe Arg Gly Asp Asn Arg Thr Pro Glu Gln Ile
115 120 125
Val Ala Ala Gly Gly Phe Tyr Pro Trp Ser Lys Gln Asp Val Gly Lys
130 135 140
Ile Lys Lys Glu Leu Ile Asp Glu Phe Ile Glu Ile Gly Pro Ser Ala
145 150 155 160
His Met Met Gly His Val Arg Ser Pro Asn Lys Asn Tyr Val Ser Thr
165 170 175
Gly Met Asn Met Asp Ser Gly Gly Phe Gly Glu Gln Ser Asn Tyr Leu
180 185 190
Tyr Lys Met Glu Ile Pro Gly Leu Lys Pro Gln Asp Met Asn Glu Arg
195 200 205
Thr Leu Gly Glu Lys Ile Arg Gln Asp Lys Arg Gly Ile Asn Tyr Pro
210 215 220
His Phe Leu Met Ser His Leu Thr Leu Ala Glu Ser Glu Phe Val Ala
225 230 235 240
Met Ile Pro Ala Arg Ser Glu Glu Leu Thr Phe Ile Thr Pro Ile Pro
245 250 255
Leu Ser Tyr Ile Thr Ser Tyr Arg Lys Arg Gly Thr Asn Thr Trp Leu
260 265 270
Pro Met Pro Leu Lys Lys
275
<210> 4
<211> 1633
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 4
Met Ser Asn Tyr Glu Tyr Asp Ile Val Thr Gln His Asp Thr Tyr Gln
1 5 10 15
Ile Lys Asp Asn Glu Tyr Thr Val Val Asn Gly Lys Tyr Trp Gln Tyr
20 25 30
Glu Gln Glu Gly Asn Lys Asn Asn Asn Lys Val Ser Ile Ser Leu Met
35 40 45
Lys Glu Asn Gln Asn Asp Pro Val Trp Ile Thr Ser Asp Ile Lys Glu
50 55 60
Ile Ser Leu Tyr Ile Ile Glu Asn Leu Phe Ser Tyr His Lys Phe Ser
65 70 75 80
Ala Glu Leu Gln His Thr Leu Lys Asn Ala Val Lys Ala Val Phe Asn
85 90 95
Glu Tyr Ser Glu Ile Lys Tyr Ser Glu Leu Leu His Asn Ile Asn Asn
100 105 110
Ile Phe Asn Leu Phe Phe Ile Lys Ile Tyr Asn Thr Ser Asp Ile Asp
115 120 125
Thr Ala Ile Asn Ile Leu Thr Ala Lys Ile Glu Ile Tyr Asp Lys Leu
130 135 140
Glu Lys Ile Asn Gln Asp Lys Thr Asp Ser Asn Asn Thr Asn Val Asp
145 150 155 160
Ile Trp Glu Glu Leu Gly Ile Asn Ala Glu Glu Pro Leu Leu Lys Ile
165 170 175
Tyr Arg Gln Ala Phe Ser Thr Gly Asp Ile Asp Asp Glu Val Tyr Ser
180 185 190
Asp Ala Leu Leu Thr Phe Met Ser Asp Gly Asn Leu Glu Leu Gly Asp
195 200 205
Lys Glu Lys Ser Asp Tyr Asn Gln Arg Ile Lys Asp Lys Thr Asp Leu
210 215 220
Phe Glu Ser Tyr Lys Lys Gly Ile Glu Lys Val Ala Ser Leu Ile Thr
225 230 235 240
Thr Asn Asn Ile Asn Pro Gly Ile Pro Ile Thr Tyr Pro Glu Thr Glu
245 250 255
Lys Ser Ile Asn Ile Gly Asp Asp Leu Leu Leu Ala Gln Leu Ala Lys
260 265 270
Glu Glu Ile Ala Leu Lys Lys Gln Asn Arg Thr Glu Tyr Ser Gln Gln
275 280 285
Asp Ile Phe Glu Leu Gln Thr Leu Gln Ala Ala Lys Tyr His Leu Leu
290 295 300
Ile Leu Ser Ser Leu Gly Ala Leu Leu Tyr Gln Ile Ala Pro Asn Val
305 310 315 320
Glu Lys Met Thr Lys Gly His Gly Asp Tyr Arg Asp Ile Ile Phe Ser
325 330 335
Gln Glu Gln Ala Glu Ser Leu Phe Lys Lys His Asn Ile Gln Tyr Asp
340 345 350
Thr Asn His Val Leu Ser Gln Glu Ser Lys His Ile Glu Met Glu Gly
355 360 365
Cys Ile Ile Leu Thr Ala Ala Ile Ile Tyr Arg Met Arg Lys Glu Asn
370 375 380
Ala Thr Val Glu Gln Ala Leu Asn Tyr Ser Thr Leu Glu Thr Ile Lys
385 390 395 400
Leu Phe Glu Asn Asp Lys Lys Lys Leu Asn Pro Phe Asn Thr Asn Asn
405 410 415
Val Lys Pro Ala Gly Tyr Phe Ser Phe Ile Asp Phe Lys Lys Arg Asp
420 425 430
Lys Phe Asp Ser Gln Tyr Asn Phe Asn Glu Gln Phe Asn Val Tyr Lys
435 440 445
Asn Lys Tyr Ser His Tyr Glu Ser Ile Ser Phe Ser Lys Leu Ile Leu
450 455 460
Ser Ser Pro Ala Ala Gln Leu Thr Ala Glu Glu Ile Val Asn Pro Pro
465 470 475 480
Glu Glu Ala Phe Leu Tyr Ser Val Glu Gln Gly Met Gly Asn Val Ala
485 490 495
Met Ile Lys Met Tyr Gln Gly Asn Trp Leu Val Ile Ser Thr Ile Gln
500 505 510
Gly Gly Val Lys Ala Lys Lys Tyr Ser Arg Gln Gln Val Asp Ser Asn
515 520 525
Pro Thr Leu Arg Ala Met Ser Lys Pro Asn Ala Leu Phe Leu Ile Glu
530 535 540
Arg Lys Met Glu Thr Gly Met Gly Ile Leu Met Pro Asn Met Met Val
545 550 555 560
Asn Thr Gly Lys Arg Leu Phe Pro Thr Gly Tyr Glu Arg Ala Lys Thr
565 570 575
Leu Ser Gly Phe Ala Glu Thr Ser Arg Tyr Lys Asn Ser Tyr Asn Ala
580 585 590
Phe Trp Asn Asp Tyr Tyr Gly Ile Thr Ser Gly Met Asn Val Gly Ile
595 600 605
Ser Phe Thr Gly Ser Pro Lys Phe Asn Phe Tyr Lys Glu Glu Asn Leu
610 615 620
Leu Ser Val Thr Ala Thr Ile Ile Gln Gln Gly Leu Asn Asp Ile Ala
625 630 635 640
Ile Lys Ser Lys Gln Ala Leu Asp Ile Thr Ser Gly Trp His Ile Ala
645 650 655
Ala Thr Ile Leu Ile Pro Phe Tyr Asn Val Ile Tyr Lys Ser Thr Thr
660 665 670
Asp Ser Glu Tyr Glu Leu Thr Gly Glu Asp Ile Gly Ser Ile Val Phe
675 680 685
Asp Thr Ala Asn Val Leu Leu Val Val Ala Thr Leu Gly Met Ser Leu
690 695 700
Thr Glu Ser Met Ala Ala Lys Val Thr Gln Thr Thr Leu Arg Leu Arg
705 710 715 720
Gln Ala Gly Leu Thr Gly Arg Ala Leu Ile Thr Ala Val Val Arg Thr
725 730 735
Leu Pro Glu His Gly Ile Ile Thr Leu Arg Gln Ser Ser Gly Ile Ile
740 745 750
Leu Gly Gly Leu Ile Asp Leu Ile Glu Pro Leu Pro Ile Arg Ser Thr
755 760 765
Leu Thr Leu Thr Tyr Arg Gly Val Ile Asn Ala Val Gly Ala Met Arg
770 775 780
Asn Ser Ile Lys Leu Glu Lys Ser Phe Ala Asp Ile Phe Gly Lys Ser
785 790 795 800
Thr Arg Gly Leu Gly Lys Leu Lys Asn Glu Trp Lys Val Ser Asn Leu
805 810 815
Pro Leu Glu Glu Ile Val Pro His Ser Asn Gly Gly Glu Ile Tyr Lys
820 825 830
Gly Ile Tyr Ser Ile Arg Pro Thr Asn Pro Glu Thr Ala Val Lys Gln
835 840 845
Asn Phe Tyr Ile Lys Glu Ala Gly Ala Asn Tyr Gln Val Lys Trp Asp
850 855 860
Asp Ala Asn His Thr Trp Arg Val Val Asn Pro Thr Tyr Pro Glu Gln
865 870 875 880
Phe Ser Tyr Trp Pro Ala Val Lys Leu Asp Lys Asn Gly His Trp Val
885 890 895
Thr His Ala Asp Val Ser Asn Lys Phe Leu Ile Leu Glu Gln Ser Lys
900 905 910
Arg Ile Asp Gln Glu Leu Glu Ala Ala His Ser Asn Ile Asn Asn Asp
915 920 925
Asn Ile Leu Asp Ala Phe Ile His Ile Asn Thr Ala Phe Lys Asp Cys
930 935 940
Glu Arg Tyr Asp Ile Asp Lys Leu Ser Asp Ile Thr Asp Thr Leu Thr
945 950 955 960
His Phe Phe Glu Lys Ser Leu Lys Pro Gly Asp Lys Lys Ala Ile Phe
965 970 975
Ser Thr Glu Ile Met Ser Ile Gln Gln Ala Trp Ile Arg Glu Val Ile
980 985 990
Leu Pro Leu Gln Asn Asn Ser Ser Ile Ser Ile Glu Lys Ile Asn Ala
995 1000 1005
Ile Lys Thr Glu Leu Pro Tyr Leu Leu Arg Lys Thr Phe Pro Ile Glu
1010 1015 1020
Ser Gln Leu Pro Asn Gln Leu Val Ala Asn Lys Ile Ala Leu Ala Ile
1025 1030 1035 1040
Glu Glu Ile Pro Asn Thr Arg Ile Pro Lys Tyr Thr Ser Gly Asn Ile
1045 1050 1055
Ser Lys Thr Val Gln Tyr Thr Ser Leu Leu Glu Asn Asn His Val Asp
1060 1065 1070
Ile Pro Pro Val Gly Ile Thr Ile Thr Gly Asn Asp Thr Phe Ile Asn
1075 1080 1085
Gln Val Thr Arg Val Leu Ser Glu Ile Asp Glu Ile Pro Ser Gly Asn
1090 1095 1100
Ile Val Ile Gln Glu Leu Glu Lys Gln Gly Leu Asn Ile Gln Pro Pro
1105 1110 1115 1120
Thr Met Asn Asp Ile Val Arg Glu Lys Asn Gly Gln Phe Tyr Ala Asn
1125 1130 1135
Asn Ser Ala Gly Ser His Ile Ala Phe Asp Pro Glu Asn His Leu Ile
1140 1145 1150
Gly Thr Glu Glu Lys Leu Ile Asp Glu Pro Trp Arg Thr Arg Glu Pro
1155 1160 1165
Ala Ile Ala Leu Tyr His Glu Met Leu His Ile Tyr Tyr Asn Arg Tyr
1170 1175 1180
Pro Thr Trp Phe Thr Ser Ile Asp Asn Lys Val Ile Asp Gln Lys Val
1185 1190 1195 1200
Ser Gly Gly Phe Ser Leu Leu Glu Glu Ser Arg Ile Val Gly Thr Lys
1205 1210 1215
Tyr Tyr Val Asn Asp Lys Asn Thr Leu Phe Asp Phe Asn Asp Ser Asp
1220 1225 1230
Tyr Leu Leu Glu Asn Asn Ser Ala Leu Leu Thr Glu Asn Arg Phe Arg
1235 1240 1245
Ala Glu Tyr Ala Ile Phe Lys Asn Lys Ser Glu Tyr Val Ile Arg Pro
1250 1255 1260
Tyr Ser Gly Lys Gly Asp Ser Gln Ile Pro Leu Thr Lys Thr Lys Ile
1265 1270 1275 1280
Asn Ile Asn Glu Ser His Arg Asn Val Met Gly Val Gly Ser Gly Lys
1285 1290 1295
Pro Glu Lys Met Pro Asn Glu Ser Ala Thr Asp Tyr Arg Asn Arg Val
1300 1305 1310
Arg Glu Trp Arg Lys Ala Asn Lys Gln Pro Glu Ala Asp Ile Gly Thr
1315 1320 1325
Gly Asp Met Arg Lys Thr Lys Ala Glu Ala Arg Val Lys Leu Leu Lys
1330 1335 1340
Glu Asn Tyr Pro Gln Phe Glu Pro Gln Lys Ile Glu Leu Gly Gly Ala
1345 1350 1355 1360
Phe Gln Leu Trp Thr Val Pro Asn Glu Pro Ala Asn Lys Leu Met Leu
1365 1370 1375
Ser Ser His Gly Tyr Phe Phe Ser Asp Ser Ala Ala Thr Gln Val Pro
1380 1385 1390
Ala Gly Lys Thr Ile Gln Phe Leu Gly Pro His Gly Lys Thr Leu Leu
1395 1400 1405
Glu Ala Pro Glu Asn Pro Leu Tyr Ser Pro Phe Asp Val Thr Leu Gly
1410 1415 1420
Asn Ser Gly Phe Thr Val Gln Pro Tyr Ala Thr Ile Glu Ser Gly Asn
1425 1430 1435 1440
Lys Ala Gly Leu Gly Ser Val Lys Ile Gly Asp Lys Thr Phe Thr Val
1445 1450 1455
Asn Asp Ile Gln Asn Ile Ala Thr Asp Asp Val Glu Asn Tyr Leu Leu
1460 1465 1470
Ala Thr Gly Val Glu Ala Asn Ala Ser Asn His Gly Lys Val Arg Asn
1475 1480 1485
Tyr Gly Ile Lys Tyr Tyr Glu Lys Met Pro Asp Glu Glu Val Lys Ala
1490 1495 1500
Ala Ile Trp Lys Asn Arg Ala Asp Glu Thr Ser Thr His Lys Tyr Asp
1505 1510 1515 1520
Ala Leu Leu Val Ser Pro Glu Ala Gly Asn Arg Lys Lys Leu Ser Asp
1525 1530 1535
Ile Phe Ala Leu Met Lys Thr Asp Glu Arg Met Ser Lys Tyr Asp Glu
1540 1545 1550
Ile Thr Phe Val Ala Cys Arg Glu Glu Leu Asn Arg Ile Asn Met Lys
1555 1560 1565
Ser Ile His Asp Thr Gly Leu Gly Gly Gly Tyr Glu Pro Lys Leu Glu
1570 1575 1580
Pro Thr Val Ile Leu Ser Arg Arg Arg Arg Glu Ala Thr Phe Thr Ala
1585 1590 1595 1600
Asp Gly Ala Ile Ile Tyr Ser Ile Ile Ala Val Asn Leu His His Asn
1605 1610 1615
Phe Ile Thr Glu Glu Ile Val Gly Ile Ala Pro Phe Leu Phe Ile Asn
1620 1625 1630
Asn
<210> 5
<211> 324
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 5
Met Glu His Glu Tyr Asn Glu Lys Glu Lys Gln Arg Asn Ser Ala Ile
1 5 10 15
Lys Leu Asn Asp Ala Ile Arg Asn Asn Glu Glu Asn Met Asp Met Thr
20 25 30
Ser Pro Leu Glu Leu Asn Phe Gln Asn Thr Asn Arg Lys Ser Arg Gly
35 40 45
Leu Arg Glu Arg Phe Ser Ala Thr Leu Gln Arg Asn Leu Pro Gly His
50 55 60
Ser Met Leu Asp Arg Glu Leu Thr Thr Asp Gly Gln Lys Asn Gln Glu
65 70 75 80
Ser Arg Phe Ser Pro Gly Met Ile Met Asp Arg Leu Met His Phe Gly
85 90 95
Val Arg Thr Arg Leu Gly Lys Val Arg Asn Ser Ala Ser Lys Tyr Gly
100 105 110
Gly Gln Val Thr Phe Lys Phe Ala Gln Thr Lys Gly Thr Phe Leu Asp
115 120 125
Gln Ile Met Lys His Lys Asp Thr Ser Gly Gly Val Cys Glu Ser Ile
130 135 140
Ser Ala His Trp Ile Ser Ala His Ala Lys Gly Asp Ser Ile Phe Asn
145 150 155 160
Gln Leu Tyr Val Gly Gly Lys Lys Gly Lys Phe His Ile Asp Thr Leu
165 170 175
Phe Ser Ile Lys Gln Leu Gln Met Asp Gly Tyr Leu Asp Asp Glu Gln
180 185 190
Ser Thr Met Thr Glu Tyr Trp Leu Gly Thr Gln Gly Met Gln Pro Asn
195 200 205
Ile Gln Arg Asn Asp Asp Thr Asp Glu His Ser Ser Lys Val Val Gly
210 215 220
Glu Thr Gly Asn Arg Gly Thr Lys Asp Leu Leu His Ala Ile Leu Asp
225 230 235 240
Thr Gly Asp Lys Gly Ser Gly Tyr Lys Lys Ile Ser Phe Leu Gly Lys
245 250 255
Met Ala Gly His Thr Val Ala Ala Tyr Val Asp Asp Gln Lys Gly Val
260 265 270
Thr Phe Phe Asp Pro Asn Phe Gly Glu Phe Ser Phe Pro Asp Lys Thr
275 280 285
Ser Phe Ser His Trp Phe Thr Asp Asp Phe Trp Pro Lys Ser Trp Tyr
290 295 300
Ser Leu Glu Ile Gly Leu Gly Gln Glu Phe Glu Val Phe Asn Tyr Ala
305 310 315 320
Pro Glu Ala Pro
<210> 6
<211> 287
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 6
Met Pro Asn Lys Lys Tyr Ser Glu Asn Thr His Gln Gly Lys Lys Pro
1 5 10 15
Leu Met Lys Ser Glu Ala Asn Asn Glu His Asp Ile Gln Asn Ser Ser
20 25 30
Leu Gly Ile Gly Leu Asp Leu Asn Ser Met Met Gly Asn Ser Ser Thr
35 40 45
Ser Leu Ser His Ile Gln Asp Tyr Ser Phe Trp Lys Glu Asn Ile Ser
50 55 60
Glu Tyr Tyr Lys Trp Met Val Val Val Lys Ala His Leu Lys Gln Leu
65 70 75 80
Asp Trp Thr Leu Lys Ser Met Asp Ser Pro Glu Ser Ala Gly Thr Asn
85 90 95
Ile Ala Lys Asn Thr Gly Thr Thr Ala Leu Gln Thr Leu Leu Asn Thr
100 105 110
Gly Gly Ser Ile Ala Gly Ala Ala Ile Gly Gly Ala Ile Gly Ser Ala
115 120 125
Ile Ala Pro Gly Val Gly Thr Ile Ala Gly Met Gly Ile Gly Ala Leu
130 135 140
Ala Gly Thr Gly Leu Asn Tyr Leu Asn Asp Thr Val Ile Glu Lys Leu
145 150 155 160
Asn Glu Lys Leu Glu Ile Ala Tyr Pro Tyr Pro Lys Thr Arg Asn Met
165 170 175
Ile Phe Asp Ile Asn Asn Tyr Asp Lys Asn Pro Ile Ile Lys Ala Ile
180 185 190
Lys Lys Lys Thr Asn Lys Asp Asn Leu Lys Val Thr Ala Gly Ser Ser
195 200 205
Leu Thr Ser Gln Leu Val Gly Lys Val Thr Ser Pro Ile Lys Phe Pro
210 215 220
Ala Tyr Lys Leu Ala Asp Leu Ala Ile Ala Leu Ala Gly Leu Ser Ser
225 230 235 240
Asp Lys Ala Arg His Ile Leu Asp Phe Thr Asp Ser Ile Arg Glu Val
245 250 255
Leu Asn Glu Ser His Ser Asp Ala Val Ala Phe Met Arg Lys Asn Tyr
260 265 270
Gly Asp Asn Ala Met Gly Leu Ala Gly Leu Ser Ser Arg Ile Lys
275 280 285
<210> 7
<211> 322
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 7
Met Glu Arg Glu Tyr Ser Glu Lys Glu Lys His Lys Lys Arg Pro Ile
1 5 10 15
Gln Leu Arg Asn Ser Ile Glu Gln His Glu Glu Glu Thr Ala Asn Asn
20 25 30
Ser Leu Gly Leu Gly Leu Asp Leu Asn Gln Ala Thr Asn Pro Pro Lys
35 40 45
Val Pro Lys Asp Asn Tyr Asn Glu Glu Asn Gly Asp Leu Phe Tyr Gly
50 55 60
Leu Ala Asn Gln Arg Gly Arg Tyr Ile Lys Ser Val Asn Pro Asn Phe
65 70 75 80
Asp Pro Asp Lys Ile Asn Ser Ser Pro Met Ile Ile Asp Val Tyr Asn
85 90 95
Asn Asn Val Ser Asn Thr Ile Leu Asn Lys Tyr Pro Leu Asp Lys Leu
100 105 110
Val Lys Leu Ser Gly Asn Pro Gln Lys Tyr Ala Asn Asn Ile Lys Val
115 120 125
Glu Asn Ser Leu Gln Gln Asp Val Ala Ser Ser Lys Arg Gly Trp Tyr
130 135 140
Pro Leu Trp Asn Asp Tyr Phe Lys Thr Gly Asn Glu Asn Lys Lys Phe
145 150 155 160
Asn Ile Ala Asp Ile Tyr Lys Glu Thr Arg Asn Gln Tyr Gly Ser Asp
165 170 175
Tyr Tyr His Thr Trp His Thr Pro Thr Gly Ala Ala Pro Lys Leu Leu
180 185 190
Trp Lys Arg Gly Ser Lys Leu Gly Ile Glu Met Ala Ala Ser Asn Glu
195 200 205
Lys Thr Lys Ile His Phe Val Leu Asp Gly Leu Asn Ile Gln Glu Val
210 215 220
Val Asn Lys Gln Lys Gly Ser Thr Pro Leu Glu Gln Gly Arg Gly Glu
225 230 235 240
Ser Ile Thr Ala Ser Glu Leu Arg Tyr Ala Tyr Arg Asn Arg Glu Arg
245 250 255
Leu Ala Gly Lys Ile His Phe Tyr Glu Asn Asp Gln Glu Thr Val Ala
260 265 270
Pro Trp Glu Lys Ser Pro Glu Leu Trp Gln Asn Tyr Ile Pro Lys Asn
275 280 285
Lys Asn Gln Asn Glu Ser Ser Thr Pro Gln Arg Asn Asn Gly Thr Leu
290 295 300
Tyr Arg Leu Gly Gly Pro Phe Arg Lys Leu Arg Ala Ser Leu Arg Lys
305 310 315 320
Arg Ser
<210> 8
<211> 308
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 8
Met Met Glu His Glu Tyr Ser Lys Glu Glu Glu Lys Lys Arg Gln Gln
1 5 10 15
Ser Lys Pro Asn Asn Ala Thr His Asp Glu Ser Asn Leu Pro Leu Glu
20 25 30
Leu Glu Lys His Phe Asn Ala Arg Thr Pro Ala Thr Ala His Ser Lys
35 40 45
Trp Phe Thr Tyr Glu Asn Asp Thr Glu Val Glu Leu Thr Thr Glu Arg
50 55 60
Ile Lys Glu Ile Phe Ser Asn Lys Gln Pro Lys Ile Ile Ile Ala Gly
65 70 75 80
Asp Gly His Asn Lys Pro Pro Phe Gln Tyr Ala Lys Asn Ile Pro Asp
85 90 95
Val Asn Ser Ser Phe Asp Ala Gly Thr Leu Gln Leu Tyr Ile Glu Ala
100 105 110
Thr Asp Glu Gln Ile Asn Glu Asn Asn Pro Glu Tyr Ile Pro Lys Glu
115 120 125
Phe Met Ala Lys Pro Gly Leu Phe Thr Asn Lys Asn Arg Arg Ala Glu
130 135 140
Ile Val Gly Trp Glu Asp Ser Glu Leu Ser Asn Ala Met Lys Glu Met
145 150 155 160
Phe Glu Leu Ser Asp Lys Ser Thr Arg Glu Lys Leu Thr Pro Glu Glu
165 170 175
Thr Ser Ser Phe Tyr Lys Leu His Glu Thr Ala Ile Arg His Phe Phe
180 185 190
Arg Pro Glu Phe Asn Gln Leu Arg Asp Glu Phe Phe Glu Ile Leu Ala
195 200 205
Lys Ala Gly Ser Asn Arg Glu Leu Asp Lys Ile Ala Leu Glu Met Ile
210 215 220
Gly Phe Thr Ser Gly Thr Trp Arg Asp Glu Tyr Ile Asn Pro Thr Leu
225 230 235 240
Ala Glu Lys Ile Ala Lys His Ala Ala Glu Lys Glu Asn His Thr Phe
245 250 255
Val Val Ser Ile Gly Asp Ala His Leu Ser Glu Asn Pro Met Gln Glu
260 265 270
Tyr Leu Asn Lys Arg Arg Asn Gly Gly Glu Phe Lys His Gln Ile Ile
275 280 285
Phe Thr Arg Asp Lys Arg Pro Ile Leu Pro Asp Asn Met Lys Thr Gly
290 295 300
Asn Lys Asn Ser
305
<210> 9
<211> 298
<212> PRT
<213> 光杆状菌(Photorhabdus)
<220>
<221> VARIANT
<222> (148)..(148)
<223> Xaa 可以是任何天然存在的氨基酸
<220>
<221> UNSURE
<222> (148)..(148)
<223> The 'Xaa' at location 148 stands for Gln, Arg, Pro, or Leu.
<400> 9
Met Leu Lys Tyr Ala Asn Pro Gln Ala Val Pro Thr Gln Arg Thr Lys
1 5 10 15
Asn Thr Ala Lys Lys Pro Ser Ser Ser Ser Ser Phe Asp Gly Gln Leu
20 25 30
Glu Leu Ser Asn Gly Glu Trp Ser Lys His Ser Glu Met Gly Leu Lys
35 40 45
Arg Gly Gly Leu Ile Asn Ser Ile Arg Arg Arg Ile Ala Arg Asn Gly
50 55 60
Asn Ile Gly Arg Phe Asn Glu Leu Ile Asp Ser Glu Ala Lys Lys Trp
65 70 75 80
Pro Ser Glu Pro Val Asp Lys Asn Ile His Met Ile Trp Ile Gly Thr
85 90 95
Arg Asn Ile Ser Glu Lys Asn Ile Lys Leu Ser Ile Asp Thr Ala Lys
100 105 110
Lys Asn Pro Asp Tyr Asn Thr Ser Ile Ile Tyr Asp Ser Gly Ile Ser
115 120 125
Gly His Glu Gly Ala Arg Asn Phe Met Leu Glu Lys Phe Glu Gly Ser
130 135 140
Asn Val Asn Xaa Ser Leu Ala Phe Pro Lys Gly Ile Gly Val Met Arg
145 150 155 160
Glu Tyr Ala Pro Glu Ala Gly Lys Ala Thr Ala Phe Pro Asn Thr Pro
165 170 175
Ile Ala Val Thr Lys Asn Asn Pro Ile Ile Asn Lys Thr Leu Asp Leu
180 185 190
Ala Val Gly Asn Tyr Gln Arg Gly Glu Lys Asn Val Leu Lys Leu Ala
195 200 205
Gly Pro Asp Val Phe Thr Gln Ala Leu Tyr Gln Glu Ile Pro Gly Leu
210 215 220
Asn Ser Lys Val Leu Asn Ala Gln Leu Asp Gln Phe Glu Leu Ala Lys
225 230 235 240
Arg Gln Ala Leu Gly Leu Pro Leu Glu Lys Pro Lys Ser Phe Ala Asp
245 250 255
Glu Lys Leu Thr Ser Val Glu Lys Glu Lys Ile Asn Arg Pro Tyr Gln
260 265 270
Ser Met Arg Gly Leu Ser Gly His Val Met Asn Gly Ala Asp His Ser
275 280 285
Trp Ala Val Asp Thr Glu Val Leu Gly His
290 295
<210> 10
<211> 299
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 10
Met Met Arg Glu Tyr Ser Asn Glu Asp Asp Cys Thr Lys Glu Lys Thr
1 5 10 15
Asn Leu Val Lys Ser Glu Asn Val Glu Ala Asp Asn Tyr Leu Glu Met
20 25 30
Glu His Leu Thr Tyr Leu Ala Lys Leu Ile Ser Met Thr Glu Arg Glu
35 40 45
Asn His His Leu Asn Ser Ile Lys Leu Ile Asp Asp Ile Ile Glu Leu
50 55 60
His Asn Asp Arg Lys Gly Asn Lys Leu Leu Trp Asn Asp Asn Trp Gln
65 70 75 80
Asp Lys Ile Ile Asp Arg Asp Leu Gln Ser Ile Phe Lys Lys Ile Asp
85 90 95
Glu Met Val Ser Glu Phe Gly Gly Leu Glu Ala Tyr Lys Asp Ile Val
100 105 110
Gly Glu Ser Pro Tyr Asp Pro Thr Glu Pro Val Cys Gly Tyr Ser Ala
115 120 125
Gln Asn Ile Phe Lys Leu Met Thr Glu Gly Glu Tyr Ala Val Asp Pro
130 135 140
Val Lys Met Ala Lys Thr Gly Lys Ile Asn Gly Asn Gln Phe Ala Glu
145 150 155 160
Lys Leu Glu His Leu Asn Ser Ser Asn Asn Tyr Val Ala Leu Ile Asn
165 170 175
Asp His Arg Leu Gly His Met Phe Leu Val Asp Ile Pro Ser Thr Asn
180 185 190
Arg Glu Arg Val Gly Tyr Ile Tyr Gln Ser Asp Leu Gly Asp Gly Ala
195 200 205
Leu Pro Ala Leu Lys Ile Ala Asp Trp Leu Lys Ser Arg Gly Lys Glu
210 215 220
Ser Ile Asn Val Asn Lys Leu Lys Lys Phe Leu Asn Asp Glu Phe Thr
225 230 235 240
Met Leu Pro Glu Asn Glu Gln Lys Gly Leu Ile Ala Glu Ile Phe Asp
245 250 255
Leu Asn Lys Asp Ile Asp Ser Val Lys Ser Gly Lys Ile Lys Lys Asp
260 265 270
Lys Ala Val Asp Ile Tyr Leu Arg Glu Tyr Asp Ile Asn Asp Phe Ile
275 280 285
Ser Asn Val Glu Lys Leu Lys Thr Lys Leu Ala
290 295
<210> 11
<211> 148
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 11
Met Ile Phe Lys Met Leu Asn Leu Ala Val Phe Tyr Leu Leu Gly Asn
1 5 10 15
Ile Phe His Tyr Leu Ile Cys Gln Lys Phe Ile Cys Tyr Phe Cys Ser
20 25 30
Val Leu Lys Ser Val Thr Met Phe Leu Thr Lys Val Ala Val Gln Ile
35 40 45
Ala Leu Tyr Leu Asn Ile Leu Pro Thr Met Ala Gly Ile Ala Gly Leu
50 55 60
His Ala Glu Val Gln Ala Leu Asn Asn Leu Phe Ile Ser Gly Asp Arg
65 70 75 80
Gly Thr Glu Lys Arg Glu Asn Trp Lys Tyr Ile Arg Asn Met Leu Glu
85 90 95
Ser Thr Ile Phe Thr Gln Arg Leu Thr Ala Gly Gln Ala Gly Lys Asp
100 105 110
Phe Ala Ala Cys His Asn Cys Ser Gly Ile Leu Ser Ser Pro Val Asn
115 120 125
Val Ile Thr Gly Lys Val Glu Ser Ala Gly Gly Asn Phe Phe Ile Asn
130 135 140
Ile Ile Ser Ile
145
<210> 12
<211> 170
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 12
Met Glu Arg Glu Tyr Ser Glu Lys Pro Lys Asn Leu Ser Gln Leu Ser
1 5 10 15
Arg Lys Thr Ala Ile Ser Glu Arg Arg Ala Met Phe Glu Arg Asn Ala
20 25 30
Ser Ser Asn Asn Glu Gln Pro Val Pro Gln Phe Ala Arg Ser Tyr Thr
35 40 45
Ser Asn Arg Ser Val Val Asn Ile Asn Pro Gly Arg Ser Ser Ile Ala
50 55 60
Val Val Thr Ala Asn Ser Thr Ser Pro Val Asn Ile Ser Thr Pro Ala
65 70 75 80
Ala Ala Ser Pro Asp Lys Leu Leu Pro Ser Thr Ser Cys Asp Thr Thr
85 90 95
Ser Ser Thr Leu Thr Val Gly Lys Tyr Lys Leu Glu Leu Thr Ser Gln
100 105 110
Gly Lys Val Val Val Phe Arg Gly Asp Asn Arg Thr Pro Glu Gln Ile
115 120 125
Val Ala Ala Gly Gly Phe Gly Glu Gln Ser Asn Tyr Leu Tyr Lys Met
130 135 140
Glu Ile Pro Gly Leu Lys Pro Gln Asp Met Asn Glu Arg Thr Leu Gly
145 150 155 160
Glu Lys Ile Arg Gln Asp Ser Arg Gly Asn
165 170
<210> 13
<211> 298
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 13
Met Lys Tyr Asp Pro Arg Leu Arg Thr Trp Val Glu Asp Asp Phe Asp
1 5 10 15
Tyr Glu Lys Asn Phe Lys Lys Gln Thr Asp Tyr Ile Asn Tyr Lys Asp
20 25 30
Leu Glu Lys Gln Leu Lys Glu Asn Val Asp Tyr Tyr Ala Leu Leu Asp
35 40 45
Glu Asn Glu Ala Ile Ile Phe Leu Lys Glu Leu Gly Cys Asp Ile Lys
50 55 60
Ser Phe Leu Asn Asp Thr Ala Phe Pro Val Thr Asp Val Leu Ser Asn
65 70 75 80
Phe Ala Gly Asn Ile Lys Asp Ala Leu Gly Val Phe Lys Val Ala Lys
85 90 95
Asn Phe Lys Pro Ile Asn Ile Gly Ile Phe Thr Tyr Ile Ile Asn Glu
100 105 110
Leu Lys Gly Lys Gly Ile Lys Ala Ile Glu Tyr Leu Gly Lys Asn Gly
115 120 125
Glu Arg Tyr Ile Lys Leu Thr Asp Arg Pro Gly Ile Arg Lys Tyr Leu
130 135 140
Asn Ala Thr Arg Tyr Leu Ile Asn Asn Lys Lys Ile Met Glu Val Gly
145 150 155 160
Ile Gly Ser Val Ala Met Glu Gly Ser Ile Val Lys Gly Ala Arg Phe
165 170 175
Gly Val Ile Tyr Ser Ala Ala Tyr Arg Ser Val Glu Leu Met Phe Lys
180 185 190
Ser Glu Tyr Asp Leu Thr Asn Phe Phe Val Asn Leu Ser Met Asp Met
195 200 205
Ala Lys Ile Ile Val Ala Thr Ile Ile Ala Lys Ser Thr Val Ala Ala
210 215 220
Ala Thr Ser Phe Val Val Thr Ala Ala Leu Ser Thr Thr Ala Ile Ala
225 230 235 240
Ile Gly Val Phe Ile Ile Gly Ala Leu Val Val Trp Gly Leu Met Trp
245 250 255
Leu Asp Asp Glu Phe Lys Ile Ser Glu Thr Ile Ile Arg Arg Leu Lys
260 265 270
Glu His Lys Val Lys Thr Pro Ile Ser Thr Tyr His Ser Asp Gln Ile
275 280 285
Phe Asn Ala Trp Gly Arg Tyr Tyr Arg Gly
290 295
<210> 14
<211> 328
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 14
Met Pro Asn Lys Lys His Ser Glu Asn Thr His Gln Gly Arg Lys Pro
1 5 10 15
Leu Ile Lys Ser Glu Ala Asn Asn Glu His Asp Ile Glu Asn Ser Ser
20 25 30
Leu Gly Ile Gly Leu Asp Leu Asn Ser Thr Ile Gly Asn Asn Ser Ala
35 40 45
Ser Leu Ser Gln Ile Gln Asp Tyr Ser Phe Trp Lys Glu Asn Ile Ser
50 55 60
Glu Tyr Tyr Lys Trp Met Val Val Val Lys Ala His Leu Lys Gln Leu
65 70 75 80
Asp Trp Thr Leu Lys Ser Met Asp Ser Ser Glu Ser Ala Gly Thr Asn
85 90 95
Ile Ala Lys Asn Ile Gly Thr Thr Ala Leu Gln Thr Leu Leu Asn Thr
100 105 110
Gly Gly Ser Ile Ala Gly Gly Ala Ile Gly Gly Ala Ile Gly Ser Ala
115 120 125
Ile Ala Pro Gly Val Gly Thr Ile Ala Gly Met Gly Ile Gly Ala Leu
130 135 140
Ala Gly Thr Gly Leu Asn Tyr Leu Asn Asp Thr Val Ile Glu Lys Leu
145 150 155 160
Asn Glu Lys Leu Glu Ile Ala Tyr Pro Tyr Pro Lys Thr Arg Asn Met
165 170 175
Ile Phe Asp Ile Asn Asn Tyr Asp Lys Asn Pro Ile Ile Lys Ala Ile
180 185 190
Lys Lys Lys Thr Asn Lys Asp Asn Leu Lys Val Thr Ala Gly Ser Ser
195 200 205
Leu Thr Ser Gln Leu Val Gly Lys Val Thr Ser Pro Ile Lys Phe Pro
210 215 220
Ala Tyr Lys Leu Ser Asp Leu Ala Ile Ser His Asn Arg Ala Leu Ala
225 230 235 240
Gly Leu Ser Ser Asp Lys Ala Arg His Ile Leu Asp Phe Thr Asp Ser
245 250 255
Ile Arg Glu Val Leu Asn Glu Ser His Ser Asp Ala Val Ala Phe Met
260 265 270
Arg Lys Asn Tyr Gly Asp Asn Ala Met Gly Leu Ser Gly Leu Ser Ser
275 280 285
Arg Ile Lys Gly Glu Lys Leu Thr Leu Ala Thr Leu Ala Arg Thr Arg
290 295 300
Asn Lys Ile Glu Asn Arg Ile Asn Ser Ile Asn Lys Gln Thr Leu Lys
305 310 315 320
Leu Ser Ser Lys Asn Ser Asn Glu
325
<210> 15
<211> 322
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 15
Met Glu Arg Glu Tyr Ser Glu Lys Glu Lys His Lys Lys Arg Pro Ile
1 5 10 15
Gln Leu Arg Asn Ser Ile Glu Gln His Glu Glu Glu Thr Ala Asn Asn
20 25 30
Ser Leu Gly Leu Gly Leu Asp Leu Asn Gln Ala Thr Asn Pro Pro Lys
35 40 45
Val Pro Lys Asp Asn Tyr Asn Glu Glu Asn Gly Asp Leu Phe Tyr Gly
50 55 60
Leu Ala Thr Gln Arg Gly Arg Tyr Ile Lys Ser Val Asn Pro Asn Phe
65 70 75 80
Asp Pro Asp Lys Ile Asn Ser Ser Pro Met Ile Ile Asp Val Tyr Asn
85 90 95
Asn Asn Val Ser Asn Thr Ile Leu Asn Lys Tyr Pro Leu Asp Lys Leu
100 105 110
Val Lys Leu Ser Gly Asn Pro Gln Lys Tyr Ala Asn Asn Ile Lys Val
115 120 125
Glu Asn Asn Leu Gln Gln Asp Val Ala Ser Ser Lys Arg Gly Trp Tyr
130 135 140
Pro Leu Trp Asn Asp Tyr Phe Lys Ile Gly Asn Glu Asn Lys Lys Phe
145 150 155 160
Asn Ile Ala Asp Ile Tyr Lys Glu Thr Arg Asn Gln Tyr Gly Ser Asp
165 170 175
Tyr Tyr His Thr Trp His Thr Pro Thr Gly Ala Ala Pro Lys Leu Leu
180 185 190
Trp Lys Arg Gly Ser Lys Leu Gly Ile Glu Met Ala Ala Ser Asn Glu
195 200 205
Lys Thr Lys Ile His Phe Val Leu Asp Gly Leu Asn Ile Gln Glu Val
210 215 220
Val Asn Lys Gln Lys Gly Ser Thr Pro Leu Glu Gln Gly Arg Gly Glu
225 230 235 240
Ser Ile Thr Ala Ser Glu Leu Arg Tyr Ala Tyr Arg Asn Arg Glu Arg
245 250 255
Leu Ala Gly Lys Ile His Phe Tyr Glu Asn Asp Gln Glu Thr Val Ala
260 265 270
Pro Trp Glu Lys Ser Pro Glu Leu Trp Gln Asn Tyr Ile Pro Lys Asn
275 280 285
Lys Asn Gln Asn Glu Ser Ser Thr Pro Gln Arg Asn Asn Gly Ala Leu
290 295 300
Tyr Arg Leu Gly Gly Pro Phe Arg Lys Leu Arg Ala Ser Leu Arg Lys
305 310 315 320
Arg Ser
<210> 16
<211> 308
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 16
Met Met Glu His Glu Tyr Ser Lys Glu Glu Glu Lys Lys Arg Gln Gln
1 5 10 15
Ser Lys Pro Asn Asn Ala Thr His Asp Glu Ser Asn Leu Pro Leu Glu
20 25 30
Leu Glu Lys His Ser Asn Ala Arg Thr Ser Ala Thr Ala Tyr Ser Lys
35 40 45
Trp Phe Thr Tyr Glu Asn Asp Met Glu Val Glu Leu Thr Thr Glu Arg
50 55 60
Val Arg Glu Ile Phe Ser Asn Lys Gln Pro Lys Ile Ile Ile Ala Gly
65 70 75 80
Asp Gly His Asn Lys Pro Pro Phe Gln Tyr Thr Lys Asn Ile Pro Asp
85 90 95
Val Asn Ser Ser Phe Asp Ala Gly Thr Leu Gln Leu Tyr Ile Glu Ala
100 105 110
Thr Asp Glu Gln Ile Asn Glu Asn Asn Pro Glu Tyr Ile Pro Lys Glu
115 120 125
Phe Met Ala Lys Pro Gly Leu Phe Thr Asn Lys Asn Arg Arg Ala Glu
130 135 140
Ile Val Gly Trp Glu Asp Ser Glu Leu Ser Asn Ala Met Lys Glu Met
145 150 155 160
Phe Glu Leu Ser Asp Lys Ser Thr Arg Glu Lys Leu Thr Pro Glu Glu
165 170 175
Thr Ser Ser Phe Tyr Lys Leu His Glu Thr Ala Ile Arg His Phe Phe
180 185 190
Arg Pro Glu Phe Asn Gln Leu Arg Asp Glu Phe Phe Glu Ile Leu Ala
195 200 205
Lys Ala Gly Ser Asn Arg Glu Leu Asp Lys Ile Ala Leu Glu Met Ile
210 215 220
Gly Phe Thr Ser Gly Thr Trp Arg Asp Glu Tyr Ile Asn Pro Thr Leu
225 230 235 240
Ala Glu Lys Ile Ala Lys His Ala Ala Glu Lys Glu Asn His Thr Phe
245 250 255
Val Val Ser Ile Gly Asp Ala His Leu Ser Glu Asn Pro Met Gln Glu
260 265 270
Tyr Leu Asn Lys Arg Arg Asn Gly Gly Glu Phe Lys His Gln Ile Ile
275 280 285
Phe Thr Arg Asp Lys Arg Pro Ile Leu Pro Asp Asn Met Lys Thr Gly
290 295 300
Lys Lys Asn Ser
305
<210> 17
<211> 1633
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 17
Met Ser Asn Tyr Glu Tyr Asp Ile Val Thr Gln His Asp Thr Tyr Gln
1 5 10 15
Ile Lys Asp Asn Glu Tyr Thr Val Val Asn Gly Lys Tyr Trp Gln Tyr
20 25 30
Glu Gln Glu Gly Asn Lys Asn Asn Asn Lys Ile Ser Ile Ser Leu Met
35 40 45
Lys Asp Asn Gln Asn Asp Pro Val Trp Ile Thr Ser Asp Ile Lys Glu
50 55 60
Ile Ser Leu Tyr Ile Ile Glu Asn Leu Phe Ser Tyr His Lys Phe Ser
65 70 75 80
Ala Glu Leu Gln His Thr Leu Lys Asn Ala Val Lys Ala Val Phe Asn
85 90 95
Glu Tyr Ser Glu Ile Lys Tyr Ser Glu Leu Leu His Asn Ile Asn Asn
100 105 110
Ile Phe Asn Leu Phe Phe Ile Lys Thr Tyr Asn Thr Ser Asp Ile Asn
115 120 125
Thr Ala Ile Asn Ile Leu Thr Ala Lys Ile Glu Ile Tyr Asp Lys Leu
130 135 140
Glu Lys Ile Asn Gln Asp Lys Thr Asp Leu Asn Asn Thr Lys Val Asp
145 150 155 160
Ile Trp Glu Glu Leu Gly Ile Asn Ala Glu Glu Pro Leu Leu Lys Ile
165 170 175
Tyr Arg Gln Ala Phe Ser Thr Gly Asp Ile Asp Asp Glu Val Tyr Ser
180 185 190
Asp Ala Leu Leu Thr Phe Met Ser Asp Gly Asn Leu Lys Leu Gly Asp
195 200 205
Lys Glu Lys Ser Asp Tyr Asn Gln Arg Ile Lys Asp Lys Thr Asp Leu
210 215 220
Phe Glu Ser Tyr Lys Lys Gly Ile Glu Lys Val Ala Ser Leu Ile Thr
225 230 235 240
Thr Asn Asn Ile Asn Pro Gly Ile Pro Ile Thr Tyr Pro Glu Thr Glu
245 250 255
Lys Ser Ile Asn Ile Gly Asp Asp Leu Leu Leu Ala Gln Leu Ala Lys
260 265 270
Glu Glu Ile Ala Leu Lys Lys Gln Asn Arg Thr Glu Tyr Ser Gln Gln
275 280 285
Asp Ile Phe Glu Leu Gln Thr Leu Gln Ala Ala Lys Tyr His Leu Leu
290 295 300
Ile Leu Ser Ser Leu Gly Ala Leu Leu Tyr Gln Ile Ala Pro Asn Val
305 310 315 320
Glu Lys Met Thr Lys Gly His Gly Asp Tyr Arg Asp Ile Ile Phe Ser
325 330 335
Gln Glu Gln Ala Glu Ser Leu Phe Lys Lys His Asn Ile Gln Tyr Asp
340 345 350
Thr Asn His Val Leu Ser Gln Glu Ser Lys His Ile Glu Met Glu Gly
355 360 365
Cys Ile Ile Leu Thr Ala Ala Ile Ile Tyr Arg Met Arg Lys Glu Asn
370 375 380
Ala Thr Val Glu Gln Ala Leu Asn Tyr Ser Thr Leu Glu Thr Ile Lys
385 390 395 400
Leu Phe Glu Asn Asp Lys Lys Lys Leu Asn Pro Phe Asn Thr Asn Asn
405 410 415
Val Lys Pro Ala Gly Tyr Phe Ser Phe Ile Asp Phe Lys Lys Arg Asp
420 425 430
Lys Phe Asp Ser Gln Tyr Asn Phe Asn Glu Gln Phe Asn Val Tyr Lys
435 440 445
Asn Lys Tyr Ser His Tyr Glu Ser Ile Ser Phe Ser Lys Leu Ile Leu
450 455 460
Ser Ser Pro Ala Ala Gln Leu Thr Ala Glu Glu Ile Val Asn Pro Pro
465 470 475 480
Glu Glu Thr Phe Leu Tyr Ser Val Glu Gln Gly Met Gly Asn Val Ala
485 490 495
Met Ile Lys Met Tyr Gln Gly Asn Trp Leu Val Val Ser Thr Ile Gln
500 505 510
Gly Gly Val Lys Ala Arg Lys Tyr Ser Gln Gln Gln Val Asp Ser Gln
515 520 525
Pro Thr Leu Arg Ala Met Ser Arg Pro Asn Ala Leu Phe Leu Ile Glu
530 535 540
Arg Lys Ile Met Ile Gly Ile Gly Ile Phe Met Glu Asn Gln Ile Val
545 550 555 560
Asn Thr Gly Lys Arg Leu Phe Pro Thr Gly Tyr Glu Arg Ala Lys Thr
565 570 575
Leu Ser Gly Phe Ala Glu Thr Ser Arg Tyr Lys Asn Ser Tyr Asn Ala
580 585 590
Phe Trp Asn Asp Tyr Tyr Gly Ile Thr Ser Gly Met Asn Val Gly Ile
595 600 605
Ser Phe Thr Gly Ser Pro Lys Phe Asn Phe Tyr Lys Glu Glu Asn Leu
610 615 620
Leu Ser Val Thr Ala Thr Ile Ile Gln Gln Gly Leu Asn Asp Ile Ala
625 630 635 640
Ile Lys Ser Lys Gln Ala Leu Asp Ile Thr Ser Gly Trp His Ile Ala
645 650 655
Ala Thr Ile Leu Ile Pro Phe Tyr Asn Val Ile Tyr Lys Ser Thr Thr
660 665 670
Asp Ser Glu Tyr Glu Leu Thr Gly Glu Asp Ile Gly Ser Ile Val Phe
675 680 685
Asp Thr Ala Asn Val Leu Leu Val Val Ala Thr Leu Gly Met Ser Leu
690 695 700
Thr Glu Ser Met Ala Ala Lys Val Thr Gln Thr Thr Leu Arg Leu Arg
705 710 715 720
Gln Ala Gly Leu Thr Gly Arg Ala Leu Ile Thr Ala Val Val Arg Thr
725 730 735
Leu Pro Glu His Gly Ile Ile Thr Leu Arg Gln Ser Ser Gly Ile Ile
740 745 750
Leu Gly Gly Leu Ile Asp Leu Ile Glu Pro Leu Pro Ile Arg Ser Thr
755 760 765
Leu Thr Leu Thr Tyr Arg Gly Val Ile Ser Ala Val Gly Ala Met Arg
770 775 780
Asn Ser Ile Lys Leu Glu Lys Ser Phe Ala Asp Ile Phe Gly Lys Ser
785 790 795 800
Thr Arg Gly Leu Gly Lys Leu Lys His Glu Trp Lys Val Ser Asn Leu
805 810 815
Pro Leu Glu Glu Ile Val Pro His Ser Asn Gly Gly Glu Ile Tyr Lys
820 825 830
Gly Ile Tyr Ser Ile Arg His Thr Asn Pro Glu Thr Ala Val Lys Gln
835 840 845
Asn Phe Tyr Ile Lys Glu Ala Gly Ala Asn Tyr Gln Val Lys Trp Asp
850 855 860
Asp Ala Asn His Thr Trp Arg Val Val Asn Pro Thr Tyr Pro Glu Gln
865 870 875 880
Phe Ser Tyr Trp Pro Ala Val Lys Leu Asp Lys Asn Gly His Trp Val
885 890 895
Thr His Ala Asp Ile Ser Asn Lys Phe Leu Ile Leu Glu Lys Ser Lys
900 905 910
Arg Ile Asp Gln Glu Leu Glu Ala Ala His Ser Asn Ile Asn Asn Asp
915 920 925
Asn Ile Leu Asp Ala Phe Ile His Ile Asn Thr Ala Phe Lys Asp Cys
930 935 940
Glu Arg Tyr Asp Ile Asp Lys Leu Ser Asp Ile Thr Asp Thr Leu Thr
945 950 955 960
His Phe Phe Glu Lys Ser Leu Lys Pro Gly Asp Lys Lys Ala Ile Phe
965 970 975
Ser Thr Glu Ile Met Ser Ile Gln Gln Ala Trp Ile Arg Glu Val Ile
980 985 990
Leu Pro Leu Gln Asn Asn Ser Ser Ile Ser Ile Glu Lys Ile Asn Ala
995 1000 1005
Ile Lys Thr Glu Leu Pro Tyr Leu Leu Arg Lys Thr Phe Pro Ile Glu
1010 1015 1020
Ser Gln Leu Pro Asn Gln Leu Val Ala Asn Lys Ile Ala Leu Ala Ile
1025 1030 1035 1040
Glu Glu Ile Pro Asn Thr Arg Ile Pro Lys Tyr Thr Ser Gly Asn Ile
1045 1050 1055
Ser Lys Thr Val Gln Tyr Thr Ser Leu Leu Glu Asn Asn His Val Asp
1060 1065 1070
Ile Pro Pro Val Gly Ile Thr Ile Thr Gly Asn Asp Thr Phe Ile Asn
1075 1080 1085
Gln Val Thr Arg Val Leu Ser Glu Ile Asp Glu Ile Pro Ser Gly Asn
1090 1095 1100
Ile Val Ile Gln Glu Leu Glu Lys Gln Gly Leu Asn Ile Gln Pro Pro
1105 1110 1115 1120
Thr Met Asn Asp Ile Val Arg Glu Lys Asn Gly Gln Phe Tyr Ala Asn
1125 1130 1135
Asn Ser Ala Gly Ser His Ile Ala Phe Asp Pro Glu Asn His Leu Ile
1140 1145 1150
Gly Thr Glu Glu Lys Leu Ile Asp Glu Pro Trp Arg Thr Arg Glu Pro
1155 1160 1165
Ala Ile Ala Leu Tyr His Glu Met Leu His Ile Tyr Tyr Asn Arg Tyr
1170 1175 1180
Pro Thr Trp Phe Thr Ser Ile Asp Asn Lys Val Ile Asp Gln Lys Val
1185 1190 1195 1200
Ser Gly Gly Phe Ser Leu Leu Glu Glu Ser Arg Ile Val Gly Thr Lys
1205 1210 1215
Tyr Tyr Val Asn Asp Lys Asp Thr Leu Phe Asp Phe Asn Asp Ser Asp
1220 1225 1230
Tyr Leu Leu Glu Asn Asn Ser Ala Leu Leu Thr Glu Asn Arg Phe Arg
1235 1240 1245
Ala Glu Tyr Ala Ile Phe Lys Asn Lys Ser Glu Tyr Val Ile Arg Pro
1250 1255 1260
Tyr Ser Gly Lys Gly Asp Ser Gln Ile Pro Leu Thr Lys Thr Lys Ile
1265 1270 1275 1280
Asn Ile Asn Glu Ser His Arg Asn Val Met Gly Val Gly Ser Gly Lys
1285 1290 1295
Pro Glu Lys Met Pro Asn Glu Ser Ala Thr Asp Tyr Arg Asn Arg Val
1300 1305 1310
Arg Glu Trp Arg Lys Ala Asn Lys Gln Pro Glu Ala Asp Ile Gly Thr
1315 1320 1325
Gly Asp Met Arg Lys Thr Lys Ala Glu Ala Arg Val Lys Leu Leu Lys
1330 1335 1340
Glu Asn Tyr Pro Gln Phe Glu Pro Gln Lys Ile Glu Leu Gly Gly Ala
1345 1350 1355 1360
Phe Gln Leu Trp Thr Val Pro Asn Glu Pro Ala Asn Lys Leu Met Leu
1365 1370 1375
Ser Ser His Gly Tyr Phe Phe Ser Asp Ser Ala Ala Thr Gln Val Pro
1380 1385 1390
Ala Gly Lys Thr Ile Gln Phe Leu Gly Pro His Gly Lys Thr Leu Leu
1395 1400 1405
Glu Ala Pro Glu Asn Pro Leu Asn Ser Pro Phe Asp Val Thr Leu Gly
1410 1415 1420
Asn Ser Gly Phe Thr Val Gln Pro Tyr Ala Thr Ile Glu Ser Gly Asn
1425 1430 1435 1440
Lys Ala Gly Leu Gly Ser Val Lys Ile Gly Asp Lys Thr Phe Thr Val
1445 1450 1455
Asn Asp Ile Gln Asn Ile Ala Thr Asp Asp Val Glu Asn Tyr Leu Leu
1460 1465 1470
Ala Thr Gly Val Glu Ala Asn Ala Ser Asn His Gly Lys Val Arg Asn
1475 1480 1485
Tyr Gly Ile Lys Tyr Tyr Glu Lys Met Pro Asp Glu Glu Val Lys Ala
1490 1495 1500
Ala Ile Trp Lys Asn Arg Ala Asp Glu Thr Ser Thr His Lys Tyr Asp
1505 1510 1515 1520
Ala Leu Leu Val Ser Pro Glu Ala Gly Asn Arg Lys Lys Leu Ser Asp
1525 1530 1535
Ile Phe Ala Leu Met Lys Thr Asp Glu Arg Met Ser Lys Tyr Asp Glu
1540 1545 1550
Ile Thr Phe Val Ala Cys Arg Glu Glu Leu Asn Arg Ile Asn Met Lys
1555 1560 1565
Ser Ile His Asp Thr Gly Leu Gly Gly Gly Tyr Glu Pro Lys Leu Glu
1570 1575 1580
Pro Thr Val Ile Leu Ser Arg Arg Arg Arg Glu Ala Thr Phe Thr Ala
1585 1590 1595 1600
Asp Gly Ala Ile Ile Tyr Ser Ile Ile Ala Val Asn Leu His His Asn
1605 1610 1615
Phe Ile Thr Glu Glu Ile Val Gly Ile Ala Pro Phe Leu Phe Ile Asp
1620 1625 1630
Asn
<210> 18
<211> 324
<212> PRT
<213> 光杆状菌(Photorhabdus luminescens)
<400> 18
Met Glu His Glu Tyr Asn Glu Lys Glu Lys Gln Arg Asn Ser Ala Ile
1 5 10 15
Lys Leu Asn Asp Ala Ile Arg Asn Asn Glu Glu Asn Met Asp Met Thr
20 25 30
Ser Pro Leu Glu Leu Asn Ser Gln Asn Thr Asn Arg Lys Ser Arg Gly
35 40 45
Leu Arg Glu Arg Phe Ser Ala Thr Leu Gln Arg Asn Leu Pro Gly His
50 55 60
Ser Met Leu Asp Arg Glu Leu Thr Thr Asp Gly Gln Lys Asn Gln Glu
65 70 75 80
Ser Arg Phe Ser Pro Gly Met Ile Met Asp Arg Leu Met His Phe Gly
85 90 95
Val Arg Thr Arg Leu Gly Lys Val Arg Asn Ser Ala Ser Lys Tyr Gly
100 105 110
Gly Gln Val Thr Phe Lys Phe Ala Gln Thr Lys Gly Thr Phe Leu Asp
115 120 125
Gln Ile Met Lys His Lys Asp Thr Ser Gly Gly Val Cys Glu Ser Ile
130 135 140
Ser Ala His Trp Ile Ser Ala His Ala Lys Gly Asp Ser Ile Phe Asn
145 150 155 160
Gln Leu Tyr Val Gly Gly Gln Lys Gly Lys Phe His Ile Asp Thr Leu
165 170 175
Phe Ser Ile Lys Gln Leu Gln Met Asp Gly Tyr Leu Asp Asp Glu Gln
180 185 190
Ser Thr Met Thr Glu Tyr Trp Leu Gly Thr Gln Gly Met Gln Pro Asn
195 200 205
Ile Gln Arg Asn Asp Asp Thr Asp Glu His Ser Ser Lys Val Val Gly
210 215 220
Glu Thr Gly Thr Lys Gly Thr Lys Asp Leu Leu His Ala Ile Leu Asp
225 230 235 240
Thr Gly Asp Lys Gly Ser Gly Tyr Lys Lys Ile Ser Phe Leu Gly Lys
245 250 255
Met Ala Gly His Thr Val Ala Ala Tyr Val Asp Asp Gln Lys Gly Val
260 265 270
Thr Phe Phe Asp Pro Asn Phe Gly Glu Phe Ser Phe Pro Asp Lys Thr
275 280 285
Ser Phe Ser His Trp Phe Thr Asp Asp Phe Trp Pro Lys Ser Trp Tyr
290 295 300
Ser Leu Glu Ile Gly Leu Gly Gln Glu Phe Glu Val Phe Asn Tyr Ala
305 310 315 320
Pro Lys Glu Pro
<210> 19
<211> 293
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 19
Met Phe Lys Tyr Asp Thr Ser Glu Lys Met Ala Lys Phe Gly Lys Gly
1 5 10 15
Lys Thr Ser Asp Gly Met Leu Leu Asp Thr Leu Tyr Leu Glu Ile Pro
20 25 30
Asp Glu Lys Ala Val Met Ser Ala Tyr Lys Ser Gln Ile Leu Asp Glu
35 40 45
Leu Arg Asn Phe Ser Glu Lys Thr His Ser Phe Phe Ser Gly Lys Lys
50 55 60
Pro Leu Tyr Ser Lys Lys Tyr Leu Ala Asn Leu Ala Ala His Ala Gly
65 70 75 80
Tyr Val His Val Thr Asp Tyr Asn Ser Ile Gly Asn Tyr Lys Asp Gly
85 90 95
Phe Val Asn Phe Lys Asp Asn Ser Arg Asn Leu Ala Glu Gly Lys Leu
100 105 110
Phe Pro Gly Ile Arg Leu Ile Lys Arg Pro Lys Leu Ser Ile Val Arg
115 120 125
Asp Lys Glu Thr Glu Arg Trp Lys Lys Gln Glu Ser Asp Glu Ala Asp
130 135 140
Ala Tyr Glu Ile Thr Asp Ile Glu Ser Phe Ile Ser Gly Val Arg Asp
145 150 155 160
Met Tyr Ser Arg Ala Asn Val Asp Leu His Pro Val Ile Glu Ser Leu
165 170 175
Ile Arg Asn His Ile Val Asn Asn Asp His Val Leu Pro Thr Met Ala
180 185 190
Gly Ile Ala Gly Leu His Ala Glu Val Gln Ala Leu Asn Asn Leu Leu
195 200 205
Ile Leu Ala Asp Gly Arg Ala Gly Lys Ile Val Gly Gly Arg Lys Ile
210 215 220
Glu Glu Tyr Met Gln Asp Met Leu Lys Ser Phe Ile Phe Thr Gln Arg
225 230 235 240
Leu Thr Thr Lys Gln Ala Gly Asn Asp Phe Ala Ala Cys His Asn Cys
245 250 255
Ser Gly Ile Leu Ser Val Pro Ala Asn Val Ile Thr Gly Lys Val Ala
260 265 270
Ser Ala Gly Ser Asn Phe Ser Leu Ile Leu Ser Arg Tyr Lys Asn Ser
275 280 285
Gln Glu Ser Pro Ile
290
<210> 20
<211> 340
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 20
Met Leu Lys His Ala Asn Pro Gln Thr Val Ser Thr Gln Arg Thr Lys
1 5 10 15
Ser Thr Ala Lys Lys Pro Ser Ser Ser Ser Ser Phe Asp Arg Gln Phe
20 25 30
Glu Leu Ser Asn Ser Glu Asn Gln Pro Gly Glu Gly Asn Lys Asp Trp
35 40 45
Thr Ile Lys Gly Trp Arg Gln Arg Phe Ala Asp Arg Ser Leu Asn Lys
50 55 60
Gly His Ile Ser Pro Leu Met Asn Lys Gly Leu Leu Val Gly Ser Glu
65 70 75 80
Glu Ala Leu Ile Asn Val Pro Val Val Ala His Arg Tyr Asp Ser Ser
85 90 95
His Gln Leu Thr Asp Ala Gly Pro Leu Lys Ala Asp Ser His Ser Asn
100 105 110
Asn Leu Asp Pro Phe Tyr Gly Val Val Thr Gly Phe Arg Gly Asp Gln
115 120 125
Val Thr Ser Ser Glu Ser Gly Ser Gly Ser Ile Gly Gly His Trp Gly
130 135 140
Lys Asn Thr Leu Asp Ser Asn Ile Thr Gly Ile Asn Val Val Asn Gly
145 150 155 160
Ala Ser Gly Thr Val Gly Ile Arg Ile Ala Leu Lys Asp Ile Gln His
165 170 175
Gly Ala Pro Val Ile Val Thr Ser Gly Ala Leu Ser Gly Cys Thr Met
180 185 190
Val Tyr Ala Val Lys Asn Gly Tyr Phe Phe Ala Tyr His Thr Gly Gln
195 200 205
Lys Pro Gly Asp Lys Glu Trp Lys Thr Gly Arg Gln Gly Val Val Ala
210 215 220
Thr Tyr Arg Ser His Gln Ala Leu Ser Pro Asp Ser Glu Pro Met Ala
225 230 235 240
Val Gly Glu Gln Asn Asn Asp Leu Val Asn Ile Phe Ala Ser Tyr Asp
245 250 255
Gln Gly Ile Ile Thr Tyr Met Gly Lys Pro Gly Val Ile Ile Asp Asn
260 265 270
Thr Ala Glu Asn Val Gly Val Phe Asn Tyr Asp Glu Val Lys Leu Glu
275 280 285
Lys Pro Asp Ile Arg Ala Gly Tyr Ser Tyr Ala Leu Leu Ala Lys Asp
290 295 300
Asp Lys Gly Lys Val Asn Val Lys Val Leu Ser Glu Asp Val Ile Val
305 310 315 320
Pro Leu Gly Asn Lys Gly Lys Thr Ile Lys Ala Ile Asn Ser Leu Lys
325 330 335
Lys Arg Leu Leu
340
<210> 21
<211> 336
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 21
Met Pro Arg Tyr Ala Asn Tyr Gln Ile Asn Pro Lys Gln Asn Thr Lys
1 5 10 15
Asn Ser His Gly Lys Ser Ser Ser Ser Asn Phe Ser Ser Gly Tyr Phe
20 25 30
Ser Ser Ser Asn Asn Ser Leu Asp Asp Ser Leu Ile Arg Gln Gln Val
35 40 45
Lys Arg Glu Phe Ile Trp Glu Gly His Met Lys Glu Ile Glu Glu Ala
50 55 60
Ser Arg Leu Gly Asn Phe Ala Val Ser Phe Arg Ala Ala Gly Gly Pro
65 70 75 80
Thr Leu Arg Ala Leu Gly Lys Gly Ala Ala Ala Lys Gly His Asp Ile
85 90 95
Leu Glu Lys Thr Ile Lys Pro Gly Ser Ile Asn Lys Ala Tyr Pro Lys
100 105 110
Asp Glu Ala Ser Asn Val Ile Lys Lys Val Gln Glu Ala Gly Ile Glu
115 120 125
Gly Tyr Val Gly His Trp Asp Lys Lys Thr Gly Arg Leu Leu Gly Ile
130 135 140
Tyr Met Ser Ser Gly His Gly Leu Ser Asp Glu Gln Val Asn Gly Lys
145 150 155 160
Ile Tyr Pro Ile Asp Leu Asn Asn Leu Glu Ala Ser Leu Ser Ala Leu
165 170 175
Lys Thr Lys Glu Asn Trp Ala Ala Leu Pro Phe Thr Gly Asp Tyr Asp
180 185 190
Met His Asp Met Ile Ser Phe Thr Gly Gln Pro His Ser Val Pro Ser
195 200 205
Asn Ser Ser Glu Glu Arg Lys Ile Ile Asp Arg Ile Asn Arg Leu Val
210 215 220
Ala Arg Ser Asp Pro Asn Arg Pro Phe Gly Asp Ile Glu His Asn Val
225 230 235 240
Ile Arg His Gly Ala Gln Val Ser Tyr Pro Ala Phe Ala Met Asp Lys
245 250 255
Glu Lys Glu Glu Ile Lys Lys His Gly Gly Ile Val Lys Ala Val Ala
260 265 270
Glu Pro Gly Glu Phe Pro Val Ala Ile Val Ser Lys Gly Lys Trp Thr
275 280 285
Ile Ala Asn Asn Ile Asp Glu Leu Asn Gln Phe Tyr Asn Ser Ile Gly
290 295 300
Ala Lys Met Lys Val Ser Trp Lys Pro Gly Ala Glu Asn Pro Gly Phe
305 310 315 320
Val Ser Asn Pro Gln Arg Pro Gly Met Ala Arg Phe Ser Arg Lys Arg
325 330 335
<210> 22
<211> 299
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 22
Met Met Arg Glu Tyr Ser Lys Glu Asp Asp Cys Val Lys Glu Lys Thr
1 5 10 15
Asn Leu Ala Glu Ser Glu Asn Val Glu Ala Asp Asn Tyr Leu Glu Met
20 25 30
Asp Cys Leu Asn Tyr Leu Ala Lys Leu Asn Gly Met Pro Glu Arg Lys
35 40 45
Asp His Ser Leu Asn Ser Thr Lys Leu Ile Asp Asp Ile Ile Lys Leu
50 55 60
His Asn Asp Arg Lys Gly Asn Lys Leu Leu Trp Asn Asp Asn Trp Gln
65 70 75 80
Asp Lys Ile Ile Asp Arg Asp Leu Glu Ser Ile Phe Lys Lys Ile Asp
85 90 95
Glu Met Val Ser Glu Phe Gly Gly Ile Glu Ile Tyr Lys Asp Ile Val
100 105 110
Gly Glu Asn Pro Tyr Asp Pro Thr Glu Pro Val Cys Gly Tyr Ser Ala
115 120 125
Gln Asn Ile Phe Lys Leu Met Thr Glu Gly Glu His Ala Val Asp Pro
130 135 140
Val Lys Met Ala Gln Thr Gly Lys Ile Asn Gly Asn Glu Phe Ala Glu
145 150 155 160
Lys Leu Glu Gln Leu Asn Ser Ser Asn Asn Tyr Val Ala Leu Ile Asn
165 170 175
Asp His Arg Leu Gly His Met Phe Leu Val Asp Ile Pro Ser Thr Asn
180 185 190
Arg Glu Lys Val Gly Tyr Ile Tyr Gln Ser Asp Leu Gly Asp Gly Ala
195 200 205
Leu Pro Ala Leu Lys Ile Ala Asp Trp Leu Lys Ser Arg Gly Lys Glu
210 215 220
Ser Ile Asn Val Asn Lys Leu Lys Lys Phe Leu Ser Asn Glu Phe Thr
225 230 235 240
Met Leu Ser Glu Ser Glu Gln Lys Glu Leu Ile Ala Glu Ile Phe Asp
245 250 255
Ile Asn Lys Asp Ile Ala Asn Val Lys Leu Gly Lys Ile Lys Lys Asp
260 265 270
Lys Ala Val Asp Val Tyr Leu Arg Glu Tyr Asp Leu Asn Asp Phe Ile
275 280 285
Ser Asn Ile Glu Lys Leu Lys Thr Lys Leu Val
290 295
<210> 23
<211> 327
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 23
Met Pro Ile Ile Gly His Lys Glu Asp Leu Ile Arg Thr Glu Arg Ser
1 5 10 15
Ser Val Asp Leu Thr Arg Ser Ser Asn Asn Arg Gln Thr Asp Asn Leu
20 25 30
Glu Leu Asn Ile Pro Gln His Lys Arg Asp Asn Lys Asp Ile Glu His
35 40 45
Ala Val Ile Tyr Gly Phe Ser Gln His Arg Gly Pro Glu Met Gln Lys
50 55 60
Ala Phe Ala Asp Asn Lys Asn Pro Val Thr Ile Asp Glu Tyr Asn Ala
65 70 75 80
Gly Leu Gly Ile Met Gly Glu Leu Ser Leu Ser Asp Tyr Phe Arg Ile
85 90 95
Ser Gln Asp Leu Lys Glu Asn Arg Leu Pro Glu Leu Asn Glu Lys Asn
100 105 110
Ile Gln Asn His Ser Leu Lys Tyr Phe Asp Ala Met Gly Val Asn Met
115 120 125
Lys Ser Ala Asp Pro Asn Val Lys Glu Glu Ala Lys Glu Gln Gln Arg
130 135 140
Ala Tyr Thr Arg Ser Trp Gly Phe Tyr Met Met Glu Asn Lys Glu Lys
145 150 155 160
Leu Asp Ile Gln Ser Lys Ile Asn Asn Leu Ile Pro Lys Lys Lys Ser
165 170 175
Phe Phe Ser Lys Ser Pro Gly Glu Asp Glu Tyr Lys Lys Leu Asp Glu
180 185 190
Phe Ile Leu Lys Asn Ser Asn Gly Ser Asn Leu Thr Ile Pro Lys Gln
195 200 205
Arg Lys Ile Leu Met Lys Phe Ala Ser Ala Lys Asn Ala Val Asp Val
210 215 220
Thr Lys Asn Leu Ser Gly Glu Glu Gln Thr Trp Leu Lys Asp Ile Ile
225 230 235 240
Ala Thr Ala Phe Phe Arg Gln Thr Ser Lys Leu Gly Met Ser Trp Phe
245 250 255
Ile Glu Gln Leu Ala Ser Pro Asp Phe Arg Phe Val Ile Val Gly Phe
260 265 270
Asn Gly Glu Glu Leu Thr Thr Asp Gln Ile Arg Ser Asn Lys Pro Trp
275 280 285
Lys His Gly Asn Arg Arg Lys Glu Gly Ala Ser Glu Tyr Ala Glu Pro
290 295 300
Ile Thr Phe Ser Glu Ile Arg His Ala His Arg Lys Gly Tyr Asp Ser
305 310 315 320
Lys Ile Asn Phe Ile Lys Lys
325
<210> 24
<211> 926
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 24
Met Ile Ser Thr Phe Asp Pro Ala Ile Cys Ala Gly Thr Pro Thr Val
1 5 10 15
Thr Val Leu Asp Asn Arg Asn Leu Thr Val Arg Glu Ile Val Phe His
20 25 30
Arg Ala Lys Ala Gly Gly Asp Thr Asp Thr Leu Ile Thr Arg His Gln
35 40 45
Tyr Asp Leu Arg Gly Asn Leu Thr Gln Ser Leu Asp Pro Arg Leu Tyr
50 55 60
Asp Leu Met Gln Lys Asp Asn Thr Val Gln Pro Asn Phe Tyr Trp Gln
65 70 75 80
His Asp Leu Leu Gly Arg Val Leu His Thr Val Ser Ile Asp Ala Gly
85 90 95
Gly Thr Val Thr Leu Ser Asp Ile Glu Asp Arg Pro Ala Leu Asn Val
100 105 110
Asn Ala Met Gly Val Val Lys Thr Trp Gln Tyr Glu Ala Asn Ser Leu
115 120 125
Pro Gly Arg Leu Leu Ser Val Ser Glu Gln Ser Ala Asn Glu Ala Val
130 135 140
Pro Arg Val Ile Glu His Phe Ile Trp Ala Gly Asn Ser Gln Ala Glu
145 150 155 160
Lys Asp Leu Asn Leu Ala Gly Gln Tyr Met Arg His Tyr Asp Thr Ala
165 170 175
Gly Leu Asp Gln Leu Asn Ser Leu Ser Leu Thr Gly Ala His Leu Ser
180 185 190
Gln Ser Leu Gln Leu Leu Lys Asp Asp Gln Met Pro Asp Trp Ala Gly
195 200 205
Asp Asn Glu Ser Val Trp Gln Asn Lys Leu Lys Asn Glu Val His Thr
210 215 220
Thr Gln Ser Thr Thr Asp Ala Thr Gly Ala Pro Leu Thr Gln Thr Asp
225 230 235 240
Ala Lys Glu Asn Met Gln Arg Leu Ala Tyr Asn Val Thr Gly Gln Leu
245 250 255
Lys Ser Ser Trp Leu Thr Leu Asn Gly Gln Leu Glu Gln Ile Ile Val
260 265 270
Lys Ser Leu Ala Tyr Ser Glu Ser Gly Gln Lys Ile Arg Glu Glu His
275 280 285
Gly Asn Gly Val Val Thr Lys Tyr Ser Tyr Glu Pro Asp Thr Gln Arg
290 295 300
Leu Ile Asn Ile Thr Thr Gln Arg Ser Lys Gly His Val Phe Ser Glu
305 310 315 320
Lys Leu Leu Gln Asp Leu Leu Tyr Glu Tyr Asp Pro Val Gly Asn Ile
325 330 335
Val Ser Ile Leu Asn Arg Ala Glu Ala Thr His Phe Trp Arg Asn Gln
340 345 350
Lys Val Ser Pro Arg Asn Thr Tyr Thr Tyr Asp Ser Leu Tyr Gln Leu
355 360 365
Ile Gln Ser Thr Gly Arg Glu Met Ala Asp Ile Gly Gln Gln Asn Asn
370 375 380
Lys Met Pro Thr Pro Leu Val Pro Leu Ser Ser Asp Asp Lys Val Tyr
385 390 395 400
Thr Thr Tyr Thr Arg Thr Tyr Ser Tyr Asp Arg Gly Asn Asn Leu Thr
405 410 415
Lys Ile Gln His Arg Ala Pro Ala Ser His Asn Ile Tyr Thr Thr Glu
420 425 430
Ile Thr Val Ser Asn Arg Ser Asn Arg Ala Val Leu Ser His Asn Gly
435 440 445
Leu Thr Pro Arg Glu Val Asp Ala Gln Phe Asp Ala Ser Gly His Gln
450 455 460
Ile Ser Leu Pro Thr Gly Gln Asn Leu Ser Trp Asn Gln Arg Gly Glu
465 470 475 480
Leu Gln Gln Ala Thr Thr Ile Asn Arg Asp Asn Ser Ala Thr Asp Arg
485 490 495
Glu Trp Tyr Arg Tyr Asn Ala Gly Ser Ala Arg Ile Leu Lys Val Ser
500 505 510
Glu Gln Gln Thr Gly Asn Ser Thr Gln Gln Gln Gln Val Thr Tyr Leu
515 520 525
Pro Gly Leu Glu Leu Arg Thr Thr Lys Ser Gly Thr Asn Thr Thr Glu
530 535 540
Asp Leu Gln Val Ile Thr Met Val Glu Thr Glu Arg Thr Gln Val Arg
545 550 555 560
Ile Leu His Trp Ser Ala Gly Lys Pro Asn Asp Ile Ala Asn Asn Gln
565 570 575
Val Arg Tyr Ser Tyr Asp Asn Leu Ile Glu Ser Asn Val Met Glu Leu
580 585 590
Asp Thr Lys Gly Lys Ile Ile Ser Gln Glu Glu Tyr Tyr Pro Tyr Gly
595 600 605
Gly Thr Ala Ile Trp Thr Ala Arg Asn Gln Ile Glu Ala Ser Tyr Lys
610 615 620
Thr Val Arg Tyr Ser Gly Lys Glu Arg Asp Lys Thr Gly Leu Tyr Tyr
625 630 635 640
Tyr Arg His Arg Tyr Tyr Gln Pro Trp Leu Gly Arg Trp Leu Ser Ala
645 650 655
Asp Pro Ala Gly Thr Val Asp Gly Leu Asn Leu Tyr Arg Met Val Lys
660 665 670
Asn Asn Pro Ile Arg Tyr Gln Asp Glu Ser Gly Thr Asn Ala Asn Asp
675 680 685
Lys Ala Gln Ala Ile Phe Lys Glu Gly Lys Lys Ile Ala Ile Asn Gln
690 695 700
Leu Lys Ile Ala Ser Asn Phe Leu Lys Asp Ser Lys Asn Ser Glu Asn
705 710 715 720
Ala Leu Glu Ile Tyr Arg Ile Phe Phe Gly Gly His Gln Asp Ile Glu
725 730 735
Gln Leu Pro Gln Trp Lys Lys Arg Ile Asp Ser Val Ile Tyr Gly Leu
740 745 750
Asp Lys Leu Lys Thr Thr Lys His Val His Tyr Gln Gln Asp Lys Ser
755 760 765
Gly Ser Ser Ser Thr Val Ala Asp Leu Asn Val Asp Glu Tyr Lys Lys
770 775 780
Trp Ser Glu Gly Asn Lys Ser Ile Tyr Val Asn Val Tyr Ala Asp Ala
785 790 795 800
Leu Lys Arg Val Tyr Glu Asp Pro Leu Leu Gly Arg Glu His Val Ala
805 810 815
His Ile Ala Ile His Glu Leu Ser His Gly Val Leu Arg Thr Gln Asp
820 825 830
His Lys Tyr Ile Gly Val Leu Ser Ser Pro Gly Ser His Asp Leu Thr
835 840 845
Asp Leu Leu Ser Ile Leu Met Pro Pro Ala Asn Glu Gln Asp Arg Thr
850 855 860
Glu Lys Gln Arg Arg Ala Thr Gly Ala Arg Lys Ala Leu Glu Asn Ala
865 870 875 880
Asp Ser Phe Thr Leu Ser Ala Arg Tyr Leu Tyr Tyr Thr Ala Gln Asp
885 890 895
Pro Asn Phe Leu Ser Ser Leu Arg Lys Ala His Arg Asp Phe Asn Asn
900 905 910
Lys Lys Thr Asp Arg Leu Ile Ile Arg Pro Pro Glu Arg Arg
915 920 925
<210> 25
<211> 324
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 25
Met Glu Arg Glu Tyr Asn Lys Lys Glu Lys Gln Lys Lys Ser Ala Ile
1 5 10 15
Lys Leu Asp Asp Ala Val Gly Asn Asn Glu Glu Asn Met Asp Met Thr
20 25 30
Ser Pro Leu Glu Leu Asn Ser Gln Tyr Thr Asn Arg Lys Arg Pro Gly
35 40 45
Leu Arg Glu Arg Phe Ser Ala Thr Leu Gln Arg Asn Leu Pro Gly His
50 55 60
Ser Met Leu Asp Arg Glu Leu Thr Thr Asp Gly Gln Lys Asn Gln Glu
65 70 75 80
Ser Arg Phe Ser Pro Gly Met Ile Met Asp Arg Ile Met His Leu Gly
85 90 95
Val Arg Thr Arg Leu Gly Lys Val Arg Asn Ser Ala Ser Lys Tyr Gly
100 105 110
Gly Gln Val Thr Phe Lys Phe Ala Gln Thr Lys Gly Thr Phe Leu Asp
115 120 125
Gln Ile Met Lys His Lys Asp Thr Ser Gly Gly Val Cys Glu Ser Ile
130 135 140
Ser Ala His Trp Ile Ser Ala His Ala Lys Gly Glu Ser Ile Phe Asp
145 150 155 160
Gln Leu Tyr Val Gly Gly Gln Lys Gly Lys Phe His Ile Asp Thr Leu
165 170 175
Phe Ser Ile Lys Gln Leu Gln Met Asp Gly Tyr Leu Asp Asp Glu Gln
180 185 190
Ser Thr Met Thr Glu Tyr Trp Leu Gly Thr Gln Gly Ile Gln Pro Asn
195 200 205
Arg Gln Lys Asn Asp Asn Met Asn Glu His Ser Ser Lys Ile Val Gly
210 215 220
Glu Thr Gly Thr Arg Gly Thr Lys Asp Leu Leu Arg Ala Ile Leu Asp
225 230 235 240
Thr Gly Asp Lys Gly Ser Gly Tyr Lys Lys Ile Ser Phe Leu Gly Lys
245 250 255
Met Ala Gly His Thr Val Ala Ala Tyr Val Asp Asp Gln Lys Gly Val
260 265 270
Thr Phe Phe Asp Pro Asn Phe Gly Glu Phe Asn Phe Pro Asp Lys Val
275 280 285
Ser Phe Ser His Trp Phe Thr Asp Asp Phe Trp Pro Lys Ser Trp Tyr
290 295 300
Ser Leu Glu Ile Gly Leu Gly Gln Glu Phe Glu Val Phe Asn Tyr Glu
305 310 315 320
Pro Lys Glu Pro
<210> 26
<211> 304
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 26
Met Val Tyr Glu Tyr Ala Lys Thr Asn Asp Arg Lys Arg Lys Leu Ser
1 5 10 15
Thr Gln Ser Asp Asn Tyr Glu Glu Lys Ser Phe Ser Pro Val Leu Asp
20 25 30
Leu Ser Arg Asn Asn Gln Asn Thr Pro Asn Met Glu Asp Glu Tyr Glu
35 40 45
Thr Pro Gln Asn Phe Ile Asn Arg Thr Gly Arg Glu Lys Leu Phe Arg
50 55 60
Ala Ile Arg Met Val Ala Ser Asn Lys Arg Asp Pro Ile Thr Lys Asp
65 70 75 80
Gln Val Ser Val Pro Pro Asp Gly Asn Leu Phe Thr Glu Leu Lys Asp
85 90 95
Lys His Leu Asp Arg Ala Ala Glu Tyr Lys Lys Leu Lys Thr Trp Pro
100 105 110
Thr His Ala Ser Ile Ile Ala Thr Ser Pro Ser Ala Asn Thr Pro Ile
115 120 125
Ala Gln His Val Ser Gly Asp Asp Ala Leu Ser Pro Tyr Ile Ser Thr
130 135 140
Gly Asp Lys Pro Gly Ala Val Gln Asn Thr Val Arg Asn Trp Asn Gly
145 150 155 160
Ile Gly Pro Ala Ser Glu Arg Arg Leu Arg Pro Glu Lys Thr Trp Ser
165 170 175
Pro Ile Ile Glu Ile Asp Val Asn Lys Leu Pro Asp Thr Thr Lys Ile
180 185 190
Phe Asp Leu Asn Lys Pro Asn Asn Thr Phe Phe Ser Thr Thr Asn Ser
195 200 205
Asp Ile Ala Gln Asn Ala Phe Ala Asp Lys Glu Val Leu Ile Ser Pro
210 215 220
Glu Ile Pro Gly Leu Ala Ile Thr Arg Val Ile Asn Asp Pro Glu Glu
225 230 235 240
Ile Lys Gln Ile Ala Asn Leu Asn Pro Ser Gln Ser Leu Ile Glu Lys
245 250 255
Lys Asn Thr Ile Pro Glu Glu Lys Ile Ile Phe Glu Glu Lys Lys Ser
260 265 270
Val Pro Ile His Asp Ser Asp Ala Asp Ile Pro Ser Ser Ser Phe Val
275 280 285
Phe Pro Lys Arg Lys Lys Pro Arg Asn Ile Arg Ser Arg Thr Asp Ser
290 295 300
<210> 27
<211> 542
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 27
Met Val Phe Glu His Asp Lys Thr Val Glu Arg Lys Arg Lys Pro Ser
1 5 10 15
Ile Gln Leu Gly Asn Asp Lys Glu Lys Ser Ser Glu Gln Ala Leu Glu
20 25 30
Leu Pro Gln Ser Lys Gln Asn Asn Pro Leu Leu His Asp Leu Ile Thr
35 40 45
Ser Asn Asn Leu Arg Lys Glu Ala Ala Val Phe Ala Lys Gln Ile Gly
50 55 60
Pro Ser Tyr Gln Gly Ile Leu Asp Gly Leu Glu His Leu His Asn Leu
65 70 75 80
Ser Gly Asn Glu Gln Leu Thr Ala Gly Phe Glu Leu His Arg Arg Ile
85 90 95
Thr Arg Tyr Leu Glu Glu His Pro Asp Ser Lys Arg Asn Ala Ala Leu
100 105 110
Arg Arg Thr Gln Thr Gln Leu Gly Asp Leu Met Phe Thr Gly Thr Leu
115 120 125
Gln Glu Val Arg His Pro Leu Leu Glu Met Ala Glu Thr Arg Pro Ala
130 135 140
Met Ala Ser Gln Ile Tyr Gln Ile Ala Arg Asp Glu Ala Lys Gly Asn
145 150 155 160
Thr Pro Gly Leu Thr Asp Leu Met Val Arg Trp Val Lys Glu Asp Pro
165 170 175
Tyr Leu Ala Ala Lys Ser Gly Tyr Gln Gly Lys Ile Pro Asn Asp Leu
180 185 190
Pro Phe Glu Pro Lys Phe His Val Glu Leu Gly Asp Gln Phe Gly Glu
195 200 205
Phe Lys Thr Trp Leu Asp Thr Ala Gln Asn Gln Gly Leu Leu Thr His
210 215 220
Thr Arg Leu Asp Glu Gln Asn Lys Gln Val His Leu Gly Tyr Ser Tyr
225 230 235 240
Asn Glu Leu Leu Asp Met Thr Gly Gly Val Glu Ser Val Lys Met Ala
245 250 255
Val Tyr Phe Leu Lys Glu Ala Ala Lys Gln Ala Glu Pro Gly Ser Ala
260 265 270
Lys Ser Gln Glu Ala Ile Leu Leu Asn Arg Phe Ala Asn Pro Ala Tyr
275 280 285
Leu Thr Gln Leu Glu Gln Gly Arg Leu Ala Gln Met Glu Ala Ile Tyr
290 295 300
His Ser Ser His Asn Thr Asp Val Ala Ala Trp Asp Gln Gln Phe Ser
305 310 315 320
Pro Asp Ala Leu Thr Gln Phe Asn His Gln Leu Asp Asn Ser Val Asp
325 330 335
Leu Asn Ser Gln Leu Ser Phe Leu Leu Lys Asp Arg Gln Gly Leu Leu
340 345 350
Ile Gly Glu Ser His Gly Ser Asp Leu Asn Gly Leu Arg Phe Val Glu
355 360 365
Glu Gln Met Asp Ala Leu Lys Ala His Gly Val Thr Val Ile Gly Leu
370 375 380
Glu His Leu Arg Ser Asp Leu Ala Gln Pro Leu Ile Asp Lys Phe Leu
385 390 395 400
Thr Ser Glu Asn Glu Pro Met Pro Ala Glu Leu Ala Ala Met Leu Lys
405 410 415
Thr Lys His Leu Ser Val Asn Leu Phe Glu Gln Ala Arg Ser Lys Gln
420 425 430
Met Lys Ile Ile Ala Leu Asp Asn Asn Ser Thr Thr Arg Pro Ala Glu
435 440 445
Gly Glu His Ser Leu Met Tyr Arg Ala Gly Ala Ala Asn Asn Val Ala
450 455 460
Val Glu Arg Leu Gln Gln Leu Pro Ala Glu Glu Lys Phe Val Ala Ile
465 470 475 480
Tyr Gly Asn Ala His Leu Gln Ser His Glu Gly Ile Asp His Phe Leu
485 490 495
Pro Gly Ile Thr His Arg Leu Gly Leu Pro Ala Leu Lys Val Asp Glu
500 505 510
Asn Asn Arg Phe Thr Ala Gln Ala Asp Asn Ile Asn Gln Arg Lys Cys
515 520 525
Tyr Asp Asp Val Val Glu Val Ser Arg Ile Gln Leu Thr Ser
530 535 540
<210> 28
<211> 2957
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 28
Met Lys Gly Ile Glu Gly Val Ile Met Leu Ser His Asp Ile Leu Pro
1 5 10 15
Glu Lys Leu Leu Val Ser Glu Lys Lys His Glu Asn Val Gly Ser Tyr
20 25 30
Phe Ser Asp Asp Ile Gly Glu Gln Ser Glu Gln Thr Glu Val Ser His
35 40 45
Phe Asn Leu Ser Leu Asp Asp Ala Phe Asp Ile Tyr Ala Asp Ile Ser
50 55 60
Ile Glu Asn Gln Gln Glu Leu Lys Asn Lys Asp Asn Asn Thr Asn Ile
65 70 75 80
Trp Ser Ser Leu Gly Arg Gly Asp Asp Asp His Asn Leu Lys Lys Ile
85 90 95
Ile Asn Asp Ala Phe Lys Glu Lys Leu Pro Gln Leu Met Glu Tyr Arg
100 105 110
Arg Lys Gly Tyr Asn Val Ile Gly Leu Asp Lys Glu Gly Ile Lys Lys
115 120 125
Leu Glu Gly Met Leu Lys Ala Val Pro Pro Glu Ile Gln Gln Pro Thr
130 135 140
Met Lys Asn Leu Tyr Ser Ala Ala Gln Glu Leu Leu Asn Thr Leu Lys
145 150 155 160
Gln His Pro Leu Leu Pro Glu Asn Gln Asp Met Ile Gln Gln Ser Asn
165 170 175
Leu Val Ile Arg Asn Leu Ser Asp Ala Leu Glu Ala Ile Asn Ala Val
180 185 190
Ser Lys Val Asn Gln Val Glu Trp Trp Glu Glu Val His Lys Thr Asn
195 200 205
Lys Ala Gln Ser Asp Arg Leu Ile Ala Ala Thr Leu Glu Glu Leu Phe
210 215 220
Phe Lys Val Lys Asp Lys Arg Leu Pro Gly Ser Asn Asp Asp Tyr Cys
225 230 235 240
Gln Gln Glu Arg Glu Glu Thr Glu Arg Lys Ile Lys Asp Leu Leu Leu
245 250 255
Tyr Asp Gly Tyr Gln Leu Thr Ala Glu His Phe Lys Phe Gly Arg Leu
260 265 270
Arg Lys Ser Leu Leu Ala Glu Ser Arg Val Thr Arg Leu Lys Leu Ala
275 280 285
Glu Tyr Leu Glu Lys Lys Ser Val Gly Ile Leu Thr Ala Ala Arg Asp
290 295 300
Ala Lys Met Tyr Ala Met Lys Ile Leu Leu Ala Gln Thr Arg Asn Asn
305 310 315 320
Gly Phe Asn Ala Lys Asp Leu Ile Asn Ala Gly Gln Val Asn Asp Arg
325 330 335
Leu Leu Ser Phe Gln Gln Tyr Ala Arg His Ile Arg Ala Val Asp Gly
340 345 350
Glu Ile Asp Gly Ile Ile Leu Ser Asn Pro Leu Val Val Ala Cys Ile
355 360 365
Lys Glu Thr Asn Asp Glu Pro Ala His Ile Lys Ile Ala Arg Ala Ile
370 375 380
Leu Pro Val Ser Glu Glu Leu Gly Thr Val Ser Lys Val Leu Arg Glu
385 390 395 400
Thr Lys Glu Lys Val Gln Pro Ser Lys Pro Lys Glu Glu Leu Asn His
405 410 415
Pro His Gln Asp Trp Trp Asn Arg Gly Asp Glu Leu Trp Lys Tyr Ile
420 425 430
Lys Lys Thr Ser Trp Asn Ile Lys Glu Thr Ser Val His Val Thr Gln
435 440 445
Met Val Gly Tyr Glu Ala Ser Lys Thr Ala Ser Arg Ala Lys His Lys
450 455 460
Leu Lys Glu Ser Ser Tyr Ser Glu Ser Ile Asn Gly Ala Val Lys Gly
465 470 475 480
Thr Ala Leu Leu Leu Leu Asp Glu Ile Gln Gln Ala Glu Asn Arg Ile
485 490 495
Arg Gln Ile Pro Gln Phe Ala Trp Asp Val Gln Glu Ala Val Glu Gln
500 505 510
His Ser Ser Val Ile Gln Arg Thr Ala Tyr Pro Asp Glu Leu Pro Glu
515 520 525
Leu Ser Glu Leu Leu Asn Glu Gln Leu Lys His Glu Glu Ala Arg Trp
530 535 540
Gln Ala Val Lys Lys Gln Ser Arg Asp Lys Leu Gln Glu Leu Ile Ala
545 550 555 560
Pro Ile Thr Arg Leu Ala Gln Glu Lys Trp Ala Gln Asp Leu Tyr Phe
565 570 575
Gln Leu Gly Glu Glu Leu Arg Lys Glu Arg Gln Asp Arg Trp Lys Asp
580 585 590
Ile Gln Gln Phe Asp Glu Ile Met Ala Glu Ala Val Gly Gln Phe Ala
595 600 605
Glu Met Ala Arg Glu Leu Asp Ser Glu Ala Val Arg Leu Ala Glu His
610 615 620
Gly His Ser Gly Gly Lys Glu Leu Gln Glu Lys Val Ala Lys Trp Leu
625 630 635 640
Arg Asp Leu Ser Lys Leu Lys Gly Lys Val Lys Ala Gly Val Ala Lys
645 650 655
Ile Thr Gly Thr Ser Leu Asp Asn Phe Ser Arg Ser Gly Met Leu Ala
660 665 670
Arg Gly Met Ser Glu Trp Ala Glu Asp Leu Lys Gln Ser Tyr Leu Gln
675 680 685
Glu Thr Leu Gln Glu Gly Ser Ala Val Ala Ala Glu Leu Phe Glu Arg
690 695 700
Thr Leu Met Glu Val Val Glu Glu Asn Arg Thr His Phe Ala Lys Glu
705 710 715 720
Ser Asp Pro Glu Ala Glu Arg Phe Leu Lys Arg Leu Ala Leu Ala Leu
725 730 735
Lys His Ala Ala Glu Asn Thr Thr Val Tyr Pro Pro Thr Pro Glu Glu
740 745 750
Ile Leu Ala Gly Ser Arg Ser Leu Pro Glu Asp Ile Arg His Trp Ala
755 760 765
Glu Lys Lys Val Val Ser Gly Ala Ile Ser Ala Ala Phe Arg Gly Gly
770 775 780
Phe Lys Leu Val Thr Gly Thr Phe Ser Leu Pro Val Arg Val Val Ile
785 790 795 800
Arg Gly Ala Lys Thr Gly Gly Thr Leu Tyr Arg Gly Val Arg Ala Ile
805 810 815
Asn Arg Ser Val Arg Leu Gly Gln Gly Pro Ala Thr Gln Val Lys Ser
820 825 830
Lys Phe Ile Asn Gln Glu Leu Ser Lys Thr Ala Phe Arg Leu Thr Leu
835 840 845
Ser Leu Ser Pro Leu Val Ala Trp Gly Met Ala Ala Ser Ile Thr Ala
850 855 860
Gly Arg Leu Tyr Asn Glu Lys Asp Tyr Pro Glu Lys Ile Ile Lys Asn
865 870 875 880
Ile Val Ile Asp Leu Pro Glu Glu Leu Leu Trp Ile Gly Gly Tyr Ala
885 890 895
Gly Ile Asn Ala Ala Ile Arg Ala His Ala Glu Lys Ala Ile Gln Gln
900 905 910
Ala Ile Gln His Ala Leu Asp Glu Gln Ala Asp Lys Leu Ala Leu Arg
915 920 925
Ile Asn Lys Glu Ile Ala Gly Lys Ser Ala Asp Val Asn Val Glu Ile
930 935 940
Ile Pro Gln Glu Thr Ser Val Ser Pro Ala Glu Thr Ala Gln Ser Thr
945 950 955 960
Pro Glu Pro Leu Ser Asp Phe Ala Ser Thr Ser Gln Leu Thr Met Pro
965 970 975
Glu Leu Ile Asp Ile Gln Asp Asn Asn Ser Ala Gln Gln Pro Lys Val
980 985 990
Arg Arg Lys Arg Asp Val Ser Val Glu Ser Glu Ile Ser Ile Asp Asn
995 1000 1005
Leu Asn Ile Ile Asn Ala Asn Thr Arg Glu Asp Lys Val Asn Ser Glu
1010 1015 1020
Ile Lys Ser Glu Leu Arg Ser Glu Leu Lys Arg Phe Glu Asn Ser Asp
1025 1030 1035 1040
Ala Asn Ser Pro Met Ser Asp Val Glu Arg Ala Ile Phe Ile Asp Leu
1045 1050 1055
Phe Leu Tyr Lys Asn Lys Tyr Glu Val Ser Glu Ser Gln Gln Asp Tyr
1060 1065 1070
Lys Asn Thr Trp Leu Lys Phe Arg Arg Glu Leu Glu Ser Gln Glu Asn
1075 1080 1085
Lys Glu Ile Lys Glu Tyr Leu Arg Phe Arg Ser Ile Ile Glu Ala Tyr
1090 1095 1100
Glu Ile Tyr Asp Lys Lys Arg Leu Asp Asp Asp Thr Ile Pro Glu Ala
1105 1110 1115 1120
Gly Thr Ile Ile Lys Glu Val Ile Asp Phe Phe Gln Lys Leu Lys Lys
1125 1130 1135
Glu Asn Pro Ile Thr Phe Met Lys Leu Ala Glu Ala Met Val Lys Phe
1140 1145 1150
Gln Tyr Tyr Tyr Glu Glu Glu Asp Glu Asn Glu Asp Arg Tyr Phe Lys
1155 1160 1165
Met Ala Glu Ile Tyr Tyr Phe Leu Asn Lys Thr Glu Asn Glu Lys Lys
1170 1175 1180
Ser Lys Thr Phe His Leu Asp Ile Ile Asp Lys Tyr Pro Asn Glu Asn
1185 1190 1195 1200
Asn Arg Leu Leu Asp Glu Phe Phe Leu Asn Lys Asn Asn Asn Asn Pro
1205 1210 1215
Asp Leu Asp Glu Ile Ile Tyr Lys Leu Gln Ser Met Gln Glu Lys Tyr
1220 1225 1230
Arg Glu Ser Tyr Glu Met Leu Ser Lys Val Glu Asn Ile His Gln Val
1235 1240 1245
Leu Ser Asp Asp Ser Lys Asn Glu Glu Asn Ile Phe Leu Asp Asn Arg
1250 1255 1260
Ile Ile Ala Ala Gln Val Phe Asp Gly Ser Ile Asn Ile Ser Leu Gln
1265 1270 1275 1280
Asp Lys Lys Lys Trp Leu Asn Arg Tyr Asp Gln Ile Arg Asn Glu Glu
1285 1290 1295
Gly Ser Asp Gly Trp Lys Leu Met His Ile Glu Ser Ile Leu Ile Asn
1300 1305 1310
Leu Arg Arg Ile Asn Thr Ala Ile Asn Leu Thr Ala Met Lys Ser Glu
1315 1320 1325
Ser Ala Leu Leu Leu Ile Asp Lys Leu Leu Asn Phe Gln Lys Lys Ala
1330 1335 1340
Arg Glu Asn Ile Leu His Ile Ser Glu Thr Pro His Glu Asp Phe Thr
1345 1350 1355 1360
Ser Tyr Ser Gln Phe Lys Thr Arg Lys Glu Leu Gly Asn Asp Asp Ser
1365 1370 1375
Lys Tyr Tyr Ala Gln Phe Asp Asn Tyr Lys Asp Asn His Asp Ala Glu
1380 1385 1390
Lys Glu Ala Lys Glu Ile Leu Ser Gln Val Val Ala Arg Ala Ser Leu
1395 1400 1405
Ser Phe Ser Glu Leu Phe Asp Lys Val Glu Ser Ile Lys Leu Phe Ser
1410 1415 1420
Phe Val Tyr Lys Asn Arg Asp Gly Gly Ala Pro Leu Ala Ala Pro Gly
1425 1430 1435 1440
Arg Thr Val Val Ile Lys Phe Pro Gly Lys Asp Thr Gly Gly Leu Val
1445 1450 1455
Ile Ser Asn Leu Phe Leu Arg Asn His Val Lys Arg Ile Ser Thr Lys
1460 1465 1470
Glu Met Glu Asp Leu Lys Pro Leu Thr Glu Gly Met Tyr Thr Arg Ala
1475 1480 1485
Thr Gln His Arg Ser Leu Gly Ser Tyr Tyr His Ile Gly Ser Gln Ser
1490 1495 1500
Glu His Thr Asn Ala Leu Glu Ile Leu Ser Gly Met Asn Lys Glu Glu
1505 1510 1515 1520
Leu Lys Thr His Leu Lys Lys Gln Gly Ile Trp Phe Gly Glu Pro Ala
1525 1530 1535
Leu Phe Ser Asn Glu Tyr Pro Lys Gln Glu Asn Thr Gly His Leu Glu
1540 1545 1550
Asn Thr Thr Leu Lys Asn Ala Ile Ile Gly Val Ser Thr Ile Gln Asn
1555 1560 1565
Asn Ala Ala Ala Asn Tyr Leu Arg Ser Thr Met Tyr Glu Ser Thr Gly
1570 1575 1580
Trp Glu Lys Leu Gly Asp Arg Phe Ile Pro Phe Tyr Glu Ile Gly Arg
1585 1590 1595 1600
Arg Lys His Tyr Asp Arg Glu Tyr Glu Ile Asn Ser Glu Gln Leu Thr
1605 1610 1615
Leu Asp Ile Ile Thr Ser Ile Ala Ile Ala Tyr Pro Ala Ala Arg Gly
1620 1625 1630
Ile Val Ala Thr Ile Arg Ser Ser Ala Ile Pro Ser Ile Leu Lys Ser
1635 1640 1645
Gly Leu Arg Gly Ser Ala Leu Phe Lys Ser Leu Ser Leu Glu Leu Gly
1650 1655 1660
Lys Met Gly Phe Asn Ala Ser Lys Val Phe Gly Gly Ala Val Tyr Glu
1665 1670 1675 1680
Leu Ile Glu Pro Tyr Pro Ile Asn Ser His Leu Asn Arg His Asn Val
1685 1690 1695
Phe Asn Lys Val Lys Asp Thr Ala Trp Glu Phe His Thr Asp Val Gly
1700 1705 1710
Leu Lys Gly Gly Gly Leu Lys Asp Phe Ile Asp Arg Phe Thr Lys Glu
1715 1720 1725
Pro Lys Glu Ile Thr Ile Ser Gly Tyr Lys Phe Lys Arg Ile Lys Tyr
1730 1735 1740
Asn Gln Glu Asn Phe Asp Thr Met Gln Arg Met Ala Leu Asp Tyr Ala
1745 1750 1755 1760
Tyr Asn Pro Asp Ser Lys Gly Lys Ile Ala Gln Ala Gln Gln Ala Tyr
1765 1770 1775
Lys Thr Gly Lys Glu Asp Tyr Asn Ala Pro Gln Tyr Asp Asn Phe Asn
1780 1785 1790
Gly Leu Ser Leu Asp Lys Lys Ile Glu Arg Tyr Ile Ser Pro Asp Thr
1795 1800 1805
Asp Ala Thr Thr Lys Gly Val Leu Ala Gly Lys Met Asn Glu Ser Ile
1810 1815 1820
Lys Asp Ile Asn Ala Phe Gln Thr Ala Lys Asp Ala Gln Ser Trp Lys
1825 1830 1835 1840
Lys Ser Ala Asn Lys Ala Asn Lys Val Val Leu Thr Pro Gln Asn Leu
1845 1850 1855
Tyr Leu Lys Gly Lys Pro Ser Glu Cys Leu Pro Glu Ser Val Leu Met
1860 1865 1870
Gly Trp Ala Leu Gln Ser Ser Gln Asp Ala Lys Leu Ser Lys Met Leu
1875 1880 1885
Met Gly Ile Tyr Ser Ser Asn Asp Ile Thr Ser Asn Pro Leu Tyr Lys
1890 1895 1900
Ser Leu Lys Glu Leu His Ala Asn Gly Asn Ala Ser Lys Phe Asn Ala
1905 1910 1915 1920
Ser Ala Thr Ser Ile Ser Asn Ile Asn Val Ser Asn Leu Ala Thr Ser
1925 1930 1935
Glu Thr Lys Leu Phe Pro Thr Glu Ile Ser Ser Val Arg Val Asp Ala
1940 1945 1950
Pro Lys His Thr Met Leu Ile Ser Lys Ile Lys Asn Arg Glu Asn Lys
1955 1960 1965
Ile Lys Tyr Val Phe Tyr Asp Pro Asn Tyr Gly Met Ala Tyr Phe Asp
1970 1975 1980
Lys His Ser Asp Met Ala Ala Phe Phe Gln Lys Lys Met Gln Gln Tyr
1985 1990 1995 2000
Asp Phe Pro Asp Asp Ser Val Ser Phe His Pro Leu Asp Tyr Ser Asn
2005 2010 2015
Val Ser Asp Ile Lys Ile Ser Gly Arg Asn Leu Asn Glu Ile Ile Asp
2020 2025 2030
Gly Glu Ile Pro Leu Leu Tyr Lys Gln Glu Gly Val Gln Leu Glu Gly
2035 2040 2045
Ile Thr Pro Arg Asp Gly Ile Tyr Arg Val Pro Pro Lys Asn Thr Leu
2050 2055 2060
Gly Val Gln Glu Thr Lys His Tyr Ile Ile Val Asn Asn Asp Ile Tyr
2065 2070 2075 2080
Gln Val Glu Trp Asp Gln Thr Asn Asn Thr Trp Arg Val Phe Asp Pro
2085 2090 2095
Ser Asn Thr Asn Arg Ser Arg Pro Thr Val Pro Val Lys Gln Asp Thr
2100 2105 2110
Asn Gly Glu Trp Phe Lys His Ser Glu Thr Gly Leu Lys Gly Gly Gly
2115 2120 2125
Pro Ile Asp Asp Ile Arg Lys Tyr Ile Ala Arg Lys Ser Ala Ile Lys
2130 2135 2140
Ile Phe Asn Gln Ser Ile Asn Tyr Ser Ala Thr Lys Trp Pro Pro Glu
2145 2150 2155 2160
Pro Ile Asp Lys Asn Ile His Met Ile Trp Ile Gly Thr Lys Asn Ile
2165 2170 2175
Ser Glu Lys Asn Ile Lys Leu Ser Ile Asp Thr Ala Lys Lys Asn Pro
2180 2185 2190
Asp Tyr Asn Thr Ser Ile Ile Tyr Asp Ser Gly Ile Ser Gly His Glu
2195 2200 2205
Gly Ala Lys Lys Phe Met Leu Glu Lys Phe Gln Asp Ser Asn Val Asn
2210 2215 2220
Ile Ile Asp Phe Arg Lys Lys Ser Tyr Phe Ser Gln Leu Lys Gln Glu
2225 2230 2235 2240
Pro Ser Phe Ala Tyr Tyr Glu Gln Val Ile Ala Glu Asn Lys Tyr Ala
2245 2250 2255
Gln Ala Ser Asp Ile Leu Arg Leu Leu Val Leu Lys Tyr Glu Gly Gly
2260 2265 2270
Ile Tyr Lys Asp Ile Asp Asp Ile Gln Val Lys Gly Phe Gly Ser Leu
2275 2280 2285
Thr Phe Pro Lys Gly Ile Gly Val Met Arg Glu Tyr Ala Pro Glu Ala
2290 2295 2300
Gly Lys Ala Thr Ala Phe Pro Asn Thr Pro Ile Ala Val Thr Lys Asn
2305 2310 2315 2320
Asn Pro Ile Ile Asn Lys Thr Leu Asp Leu Ala Val Ser Asn Tyr Gln
2325 2330 2335
Arg Gly Glu Lys Asn Val Leu Lys Leu Ala Gly Pro Asp Val Phe Thr
2340 2345 2350
Gln Ala Leu Tyr Gln Glu Ile Pro Gly Leu Asp Ser Lys Val Leu Asn
2355 2360 2365
Ala Gln Leu Tyr Gln Leu Glu Leu Ala Lys Arg Gln Ala Leu Gly Val
2370 2375 2380
Pro Leu Glu Lys Pro Lys Asn Phe Ala Asp Glu Gln Leu Thr Ser Ala
2385 2390 2395 2400
Glu Lys Glu Lys Ile Asn Arg Pro Tyr Gln Ser Ile Arg Gly Leu Ser
2405 2410 2415
Gly Tyr Val Glu Asn Gly Ala Asp His Ser Trp Ala Val Asp Thr Asn
2420 2425 2430
Ile Pro Ser Thr Ser Thr Gln Thr Ser Thr Ile Val Thr Pro Leu Ala
2435 2440 2445
Pro Lys Thr Glu Met Leu Pro Pro Val Pro Ser Ser Ser Thr Lys Ser
2450 2455 2460
Ser Thr Ser Ala Pro Val Leu Gln Glu Lys Ile Ser Tyr Asn Leu Ala
2465 2470 2475 2480
Thr Asp Ile Asp Ala Thr Asp Tyr Leu Asn Gln Leu Lys Gln Lys Thr
2485 2490 2495
Asn Ile Asn Asn Lys Ile Ser Ser Pro Ala Gly Gln Cys Glu Ser Leu
2500 2505 2510
Met Lys Pro Val Ser Asp Phe Met Arg Glu Asn Gly Phe Thr Asp Ile
2515 2520 2525
Arg Tyr Arg Gly Met Phe Ile Trp Asn Asn Ala Thr Glu Gln Ile Pro
2530 2535 2540
Met Asn His Phe Val Val Val Gly Lys Lys Val Gly Lys Asp Tyr Val
2545 2550 2555 2560
Phe Asp Val Ser Ala His Gln Phe Glu Asn Lys Gly Met Pro Asp Leu
2565 2570 2575
Asn Gly Pro Leu Ile Leu Ala Ala Glu Asp Trp Ala Lys Lys Tyr Arg
2580 2585 2590
Gly Ala Thr Thr Arg Lys Leu Ile Tyr Tyr Ser Asp Phe Lys Asn Ala
2595 2600 2605
Ser Thr Ala Thr Asn Thr Tyr Asn Ala Leu Pro Arg Glu Leu Val Leu
2610 2615 2620
Glu Ser Met Glu Gly Lys Thr Phe Ile Thr Ser Pro Asn Trp Tyr Gln
2625 2630 2635 2640
Thr Phe Lys Arg Thr His Asn Ile His Pro Glu Val Thr Val Ser Asp
2645 2650 2655
Pro Ala Thr Phe Ser Leu Asn Tyr Ser Val Asn Pro Thr Ala Glu Asn
2660 2665 2670
Leu Ser Pro Pro Pro Pro Pro Pro Ile Pro Ser His Gly Gln Val Pro
2675 2680 2685
Lys Thr Val Thr Pro Pro Pro Pro Pro Met Arg Ser Pro Leu Ser Leu
2690 2695 2700
Ser Gln Pro Leu Glu Arg Leu Pro Ala Asn Lys Thr Lys Pro Ile Gly
2705 2710 2715 2720
Phe Asn Pro Gly Glu Asn Lys Ala Ser Phe Ser Lys Leu Glu Glu Ala
2725 2730 2735
Gly Lys His Tyr Tyr Lys Asp Asp Lys Ser Arg Gln Ala Ala Pro Val
2740 2745 2750
Asn Thr Met Ser Asp Phe Asp Asn Arg Tyr Leu Ser His Thr Thr Glu
2755 2760 2765
Ala Pro Ala Pro Ser Asn Val Ala His Leu Ala Pro Gly Asn Ile Tyr
2770 2775 2780
Asn Thr Lys Val Thr Ala Lys Gly Ala Glu Lys Pro Ala Tyr Asp Ile
2785 2790 2795 2800
Tyr Ile Ser Lys Asp Gly Glu Ser Leu Ile Thr Ser Ser Ser Tyr Lys
2805 2810 2815
Val Asp Asp Ile Thr Thr Asp Ser Lys Phe Gly Lys Pro Leu Pro Tyr
2820 2825 2830
Ser Glu Ile Met Phe Asn Ser Leu Lys Lys Ser Gly Val Asp Pro Lys
2835 2840 2845
Asn Leu Lys Arg Ser Val Gln Ala Ser Ile Glu Asn Lys Val Thr Gln
2850 2855 2860
Asp Val Ile Ser Ala Ile Gly Thr Arg Ile Gln Arg Gly Gln Val Ile
2865 2870 2875 2880
Arg Val Ser Pro Thr Glu Asn Pro Asp Ala Phe Tyr Thr Leu Leu Gly
2885 2890 2895
Thr Asp Asn Cys Lys Ala Thr Leu His Met Leu Asn Gln His Ala Glu
2900 2905 2910
Glu Phe Gly His Lys Val Val Thr Ser Ile Glu Phe Lys Gly Thr Gly
2915 2920 2925
Tyr Leu Val Met Asn Ile Gly Thr Ser Thr Gln Thr Ser Thr Ile Val
2930 2935 2940
Thr Pro Pro Pro Met Pro Gly Thr Ser Gln Leu Val Gln
2945 2950 2955
<210> 29
<211> 327
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 29
Met Pro Asn Lys Lys Tyr Ser Glu Asn Thr His Gln Gly Lys Lys Pro
1 5 10 15
Leu Ile Lys Ser Glu Ala Asn Asn Glu His Ala Ile Asp Asn Ser Pro
20 25 30
Leu Gly Ile Gly Leu Asp Leu Asn Ser Ile Leu Gly Asn Asn Ser Ala
35 40 45
Ser Leu Ser Gln Ile His Asp Tyr Ser Phe Trp Lys Glu Asn Ile Ser
50 55 60
Glu Tyr Tyr Lys Trp Met Val Val Val Lys Ala His Leu Lys Gln Leu
65 70 75 80
Asp Trp Thr Leu Lys Ser Met Asp Ser Pro Glu Ser Ala Gly Ala Asn
85 90 95
Ile Ala Lys Asn Ile Gly Thr Thr Thr Leu Gln Thr Leu Leu Asn Thr
100 105 110
Gly Gly Ser Ile Ala Gly Gly Ala Ile Gly Gly Ala Ile Gly Ser Ala
115 120 125
Ile Ala Pro Gly Val Gly Thr Ile Ala Gly Met Gly Ile Gly Ala Leu
130 135 140
Ala Gly Thr Gly Leu Asn Tyr Leu Asn Asp Thr Ala Ile Glu Lys Leu
145 150 155 160
Asn Glu Lys Leu Glu Ile Ala Tyr Pro Tyr Pro Lys Thr Arg Asn Met
165 170 175
Ile Phe Asp Ile Asn Asn Tyr Asp Lys Asn Pro Leu Ile Lys Ala Ile
180 185 190
Lys Lys Lys Thr Lys Lys Asp Asn Leu Lys Val Met Ala Gly Ser Ser
195 200 205
Leu Thr Ser Gln Leu Leu Gly Arg Ile Thr Pro Ile Lys Ile Pro Ala
210 215 220
Tyr Lys Leu Ala Asp Leu Ala Val Ser His His Arg Ala Leu Ala Gly
225 230 235 240
Leu Ser Ser Asp Lys Ala Arg His Ile Leu Asp Phe Thr Asn Ser Ile
245 250 255
Arg Glu Val Leu Asn Glu Ser His Ser Asp Ala Val Ala Phe Met Arg
260 265 270
Lys Asn Tyr Gly Asp Asn Ala Met Gly Leu Ser Gly Leu Ser Ser Lys
275 280 285
Ile Lys Gly Asp Lys Leu Thr Leu Asp Thr Leu Ala Arg Thr Arg Asn
290 295 300
Lys Ile Glu Asn Arg Ile Asn Ser Ile Asn Lys Gln Thr Leu Lys Leu
305 310 315 320
Ser Ser Lys Asn Ser Asn Glu
325
<210> 30
<211> 322
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 30
Met Glu Arg Glu Tyr Ser Glu Lys Glu Lys His Lys Lys His Pro Ile
1 5 10 15
Gln Leu Arg Asp Ala Ile Glu Gln His Ala Glu Glu Thr Ala Asn Asn
20 25 30
Ser Leu Gly Leu Gly Leu Asp Leu His Gln Ala Ile Asn Thr Pro Lys
35 40 45
Val Pro Lys Asp Asn Tyr Asn Glu Glu Asn Gly Asp Leu Phe Tyr Gly
50 55 60
Leu Ala Ala Gln Arg Gly Arg Tyr Ile Lys Ser Val Asn Pro Asn Phe
65 70 75 80
Asp Pro Asp Lys Thr Asn Ser Ser Pro Met Val Ile Asp Val Tyr Asn
85 90 95
Asn His Val Ser Asn Thr Ile Leu Asn Lys Tyr Pro Leu Asp Lys Leu
100 105 110
Gly Lys Leu Tyr Gly Asn Pro Gln Lys Tyr Ala Lys Asp Ile Lys Val
115 120 125
Thr Asn Ser Leu Gln Gln Asp Val Ala Ala Ser Lys Arg Gly Trp Tyr
130 135 140
Pro Leu Trp Asn Asp Tyr Phe Lys Ala Gly Asn Glu Asn Lys Lys Phe
145 150 155 160
Asn Ile Ala Asp Ile Tyr Lys Glu Thr Arg Asn Gln Tyr Gly Ser Asp
165 170 175
Tyr Tyr His Thr Trp His Glu Pro Thr Gly Ala Ala Pro Lys Leu Leu
180 185 190
Trp Lys Arg Gly Ser Lys Leu Gly Ile Ala Met Ala Ala Ser Asn Glu
195 200 205
Lys Thr Lys Ile His Phe Val Leu Asp Gly Leu Asn Ile Gln Glu Val
210 215 220
Val Asn Lys Gln Lys Gly Ser Thr Pro Leu Glu Gln Gly Arg Gly Glu
225 230 235 240
Ser Ile Thr Ala Ser Glu Leu Arg Tyr Ala Tyr Arg Asn Arg Glu Arg
245 250 255
Leu Ala Gly Lys Ile His Phe Tyr Glu Asn Asp Gln Glu Thr Ile Ala
260 265 270
Pro Trp Glu Lys Ser Pro Glu Leu Trp Gln Asn Tyr Ile Pro Lys Asn
275 280 285
Lys Ser Gln Asn Glu Ser Ser Thr Pro Gln Arg Asn Asn Gly Ala Leu
290 295 300
Tyr Arg Leu Gly Gly Pro Phe Arg Lys Leu Arg Ala Ser Leu Arg Lys
305 310 315 320
Arg Ser
<210> 31
<211> 297
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 31
Met Val His Glu Tyr Ser Ile Asn Asp Arg Gln Lys Arg His Ser Phe
1 5 10 15
Ser Ser Ala Asn Pro Ile Asp Pro Glu Val Thr Asn Arg Glu Asn Ser
20 25 30
Arg His Arg Phe Pro Lys Asp Asn Tyr Asn Lys Gly His Gly Asp Leu
35 40 45
Phe Tyr Gly Leu Ala Pro Glu Arg Gly Lys Tyr Ile Lys Glu Ala Asn
50 55 60
Pro Lys Phe Asp Pro Asn Asn Pro Glu Asn Ala Ala Met Ile Ile Asp
65 70 75 80
Val Tyr Asn Asp Glu Ile Ser Arg Val Ile Leu Asn Asn Asn Ala Asn
85 90 95
Lys Ile Ser Thr Asn Arg Leu Leu Asn Phe Ile Tyr Asn Phe Arg Lys
100 105 110
Asn Arg Leu Glu Asn Leu Met Lys Asn Pro Glu Lys Tyr Ala Lys Asp
115 120 125
Ile Lys Val Lys Asp Asn Leu Arg Glu Asn Ile Ser Pro Lys Lys Ile
130 135 140
Glu Lys Tyr Pro Leu Trp Asn Asp Tyr Phe Glu Ala Gly Ile Arg Asn
145 150 155 160
Lys Lys Phe Asn Ile Ala Glu Ile Phe Lys Glu Thr Ala Ser Gln Tyr
165 170 175
Asn Ser Asp Tyr Tyr His Ala Trp His Ile Gly Gly Asn Ser Ala Pro
180 185 190
Arg Leu Leu Trp Lys Arg Gly Ser Lys Leu Gly Ile Glu Ile Ala Ala
195 200 205
Ser Asn Gln Arg Thr Lys Ile His Phe Ile Leu Asp Gly Leu Lys Ile
210 215 220
Glu Asp Val Val Asn Lys Thr Lys Gly Pro Ala Pro Leu Lys Ala Gly
225 230 235 240
Pro Gly Glu Ser Ile Thr Ala Ser Glu Leu Arg Tyr Ala Tyr Arg Asn
245 250 255
Arg Ala Arg Leu Ala Gly Arg Ile His Phe Tyr Glu Asn Gly Lys Glu
260 265 270
Thr Ile Ala Pro Trp Asp Lys Asp Pro Glu Leu Trp Gln Lys Tyr Thr
275 280 285
Pro Lys Asn Arg Ser Gly Met Glu Leu
290 295
<210> 32
<211> 340
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 32
Met Leu Lys Tyr Ala Asn Pro Gln Thr Val Ala Thr Gln Arg Thr Lys
1 5 10 15
Asn Thr Ala Lys Lys Pro Pro Ser Ser Thr Ser Phe Asp Gly His Leu
20 25 30
Glu Leu Ser Asn Gly Glu Asn Gln Pro Tyr Glu Gly His Lys Ile Arg
35 40 45
Lys Ile Lys Gly Leu Arg Gln His Leu Ala Asp Arg Ser Leu Asn Lys
50 55 60
Gly His Ile Ser Pro Leu Met Asn Lys Gly Leu Leu Val Gly Ser Lys
65 70 75 80
Asp Val Ser Ile Asp Ile Pro Val Ile Ala His Arg Tyr Asp Ser Ser
85 90 95
His Gln Leu Thr Asp Ala Glu Pro Leu Lys Ala Asp Ser His Ser Asn
100 105 110
His Leu Asp Pro Phe Tyr Gly Val Ile Ala Gly Phe Arg Gly Asp Gln
115 120 125
Val Thr Ser Ser Glu Ser Gly Ser Gly Ser Ile Gly Val His Trp Gly
130 135 140
Lys Asn Thr Leu Asp Ser Asn Ile Met Gly Val Asn Val Val Asn Gly
145 150 155 160
Ala Ser Gly Thr Val Gly Ile Arg Ile Ala Leu Lys Asp Ile Gln His
165 170 175
Gly Ser Pro Val Ile Val Thr Ser Gly Ala Leu Ser Gly Cys Thr Met
180 185 190
Val Tyr Ser Val Lys Asn Gly Tyr Phe Phe Ala Tyr His Thr Gly Gln
195 200 205
Lys Pro Gly Asn Asn Glu Trp Lys Thr Gly Arg Gln Gly Val Val Ala
210 215 220
Thr Tyr Leu Ser His Gln Ala Leu Ser Pro Asp Ser Glu Pro Met Thr
225 230 235 240
Val Gly Glu Gln Asn Asn Asp Leu Val Asn Ile Phe Ala Asn Tyr Asp
245 250 255
Gln Ser Val Ile Thr Tyr Met Gly Lys Pro Gly Val Leu Ile Asp Lys
260 265 270
Met Ala Glu Asn Val Gly Val Phe Asn Tyr Asp Glu Ile Lys Pro Glu
275 280 285
Lys Pro Ala Ile Arg Ala Gly Tyr Ser Tyr Ala Leu Leu Ala Lys Asp
290 295 300
Asp Lys Gly Lys Val Asn Val Lys Val Leu Ser Glu Asp Val Ile Val
305 310 315 320
Ser Ser Gly Lys Gln Gly Asn Thr Val Lys Ala Ile Asn Ser Leu Lys
325 330 335
Lys Arg Leu Leu
340
<210> 33
<211> 336
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 33
Met Pro Arg Tyr Ala Asn Tyr Gln Ile Asn Pro Lys Gln Asn Ile Lys
1 5 10 15
Asn Ser His Gly Lys Ser Ser Ser Ser Asp Phe Ser Ser Gly Tyr Leu
20 25 30
Ser Phe Ser Asn Asn Ser Leu Asp Asp Pro Phe Ile Arg Gln Gln Val
35 40 45
Lys Arg Glu Phe Ile Trp Glu Gly His Met Lys Glu Ile Glu Glu Ala
50 55 60
Ser Arg Leu Gly Asn Phe Ala Val Ser Phe Arg Ala Ala Gly Gly Pro
65 70 75 80
Thr Leu Arg Ala Leu Gly Lys Gly Ala Ala Ala Lys Gly His Asp Ile
85 90 95
Leu Glu Lys Thr Ile Lys Pro Gly Ser Ile Asn Lys Ala Tyr Pro Lys
100 105 110
Asp Glu Ala Ser Asp Val Ile Lys Lys Val Gln Glu Ala Gly Ile Glu
115 120 125
Gly Tyr Val Gly His Trp Asp Lys Lys Thr Gly Arg Leu Leu Gly Ile
130 135 140
Tyr Met Ser Ser Gly His Gly Leu Ser Asp Glu Gln Val Asn Gly Lys
145 150 155 160
Ile Tyr Pro Ile Asp Leu Asn Asn Leu Glu Ala Ser Leu Ser Ala Leu
165 170 175
Lys Ala Lys Glu Asn Trp Ala Ala Leu Pro Phe Thr Gly Asp Tyr Asp
180 185 190
Met His Asp Met Ile Ser Phe Thr Gly Gln Pro His Ser Val Pro Ser
195 200 205
Asn Ser Ser Glu Glu Arg Lys Ile Ile Asp Arg Ile Asn Arg Leu Val
210 215 220
Ala Arg Ser Asp Ser Asn Arg Pro Phe Gly Asp Ile Glu His Asn Val
225 230 235 240
Ile Arg His Gly Ala Gln Val Ser Tyr Pro Ala Phe Ala Met Asp Lys
245 250 255
Glu Lys Glu Glu Ile Lys Lys His Gly Gly Ile Val Lys Ala Val Ala
260 265 270
Glu Pro Gly Glu Phe Pro Val Ala Ile Val Ser Lys Gly Lys Trp Thr
275 280 285
Ile Ala Asn Asn Ile Asp Glu Leu Asn Gln Phe Tyr Asn Ser Ile Gly
290 295 300
Ala Lys Met Lys Val Ser Trp Lys Pro Gly Ala Glu Asn Pro Gly Phe
305 310 315 320
Val Ser Asn Pro Gln Arg Pro Gly Met Ala Arg Phe Ser Arg Lys Arg
325 330 335
<210> 34
<211> 328
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 34
Met Pro Asn Lys Lys Tyr Ser Glu Asn Thr His Gln Gly Lys Asn Pro
1 5 10 15
Leu Met Lys Ser Gly Ala Asn Asn Glu His Asp Leu Gln Asp Ser Pro
20 25 30
Leu Gly Ile Gly Leu Asp Leu Asn Ser Met Leu Val Asn Ser Ser Thr
35 40 45
Ser Leu Ser Gln Ile Gln Asp Tyr Ser Phe Trp Lys Glu Asn Ile Ser
50 55 60
Glu Tyr Tyr Lys Trp Met Val Val Val Glu Ser His Leu Lys Gln Leu
65 70 75 80
Asp Trp Thr Leu Lys Ser Met Asp Ser Pro Glu Ser Ala Gly Thr Asn
85 90 95
Val Ala Lys Asn Met Gly Val Thr Ala Leu Gln Ser Leu Leu Asn Thr
100 105 110
Gly Ser Ser Ile Ala Gly Gly Ala Ile Gly Gly Ala Ile Gly Ser Ala
115 120 125
Ile Ala Pro Gly Val Gly Thr Ile Ala Gly Ala Gly Ile Gly Ala Leu
130 135 140
Ala Gly Thr Gly Leu Asn Tyr Leu Asn Asp Thr Ala Met Ser Lys Leu
145 150 155 160
Ser Lys Lys Leu Glu Ile Ala His Pro Tyr Pro Lys Thr Arg Asn Met
165 170 175
Ile Leu Asp Ile Asn Asn Tyr Asp Lys Asn Pro Ile Ile Lys Ala Ile
180 185 190
Lys Lys Asn Val Asn Lys Asp Asn Leu Lys Val Thr Ala Gly Ser Ser
195 200 205
Leu Thr Ser Lys Leu Val Gly Thr Val Thr Ser Pro Ile Lys Phe Pro
210 215 220
Ala Tyr Lys Phe Ala Glu Leu Ala Val Ser His His Arg Ala Leu Glu
225 230 235 240
Gly Leu Ser Asp Asp Lys Ala Arg His Ile Leu Asp Phe Thr Asn Ser
245 250 255
Ile Arg Glu Val Leu Lys Glu Ser His Ser Asp Ala Val Ala Phe Met
260 265 270
Arg Lys Asn Tyr Gly Asp Asn Ala Met Gly Leu Ser Gly Phe Ser Ser
275 280 285
Lys Ile Lys Arg Glu Lys Leu Thr Leu Asn Thr Leu Ala Lys Thr Lys
290 295 300
Asn Glu Ile Glu Asn Arg Ile Asn Ser Ile Asn Lys Gln Thr Leu Lys
305 310 315 320
Val Ser Ser Arg Ser Arg Asn Glu
325
<210> 35
<211> 324
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 35
Met Leu Ser Thr Glu Lys His Asn Lys Asp Thr Lys His Pro Arg Asn
1 5 10 15
Arg Glu Lys Lys Phe Ser Ile Gln Pro Glu Asn Ser Thr Gln Asp Asp
20 25 30
Glu Asp Ile Lys Asn Asn Ser Leu Gly Val Gly Leu Asp Leu Asp Gln
35 40 45
Met Ile Arg Asn Thr Ser Ser Thr Leu Thr Asn Ala Pro Gln Lys Pro
50 55 60
Glu Asp Gly Tyr Tyr Tyr His Ile Ser Arg Gly Asn Asn Leu Gln Ser
65 70 75 80
Phe Leu Gln Asn Gly Phe Lys Pro Gln Gly Ser Pro Gly Pro Thr Leu
85 90 95
Ser Glu Glu Asp Phe Ser Arg Arg Lys Ile Gly Ile Ile Lys Leu Ile
100 105 110
Tyr Ser Ile Ile Ala Thr Thr Ile Asn Lys Asn Arg Lys Ala Lys Lys
115 120 125
Ile Ser Lys Asp Asn Phe Leu Met Pro Gln Glu Phe Trp His Glu Phe
130 135 140
Lys Asn Phe Tyr Gln Asn Ile Pro Thr Gln Thr Asn Ile Asp Asp Gln
145 150 155 160
Leu Leu Lys Lys Ser Ile Thr Glu Ser Ile Asp Lys Leu Asp Gln Asn
165 170 175
Lys Phe Met Glu Lys His Ser Asp Arg Lys Gln Thr Ile Ile Asn Asn
180 185 190
Glu Arg Glu Ala Ile Leu Gln Gln Asp Glu Arg Ile Asn Glu Ile Ile
195 200 205
Ser Ser Arg Ala Lys Met Ile Gln Gln Arg Glu Ala Glu Asn Thr Glu
210 215 220
Gly Tyr Ile Tyr Leu Ala Pro His Lys Asn Thr Leu Leu Glu Tyr Met
225 230 235 240
Lys His Leu Gln Glu Glu Lys Asn Leu Phe Leu Ile Leu Ala Val Lys
245 250 255
Glu Asp Ile Phe Thr Glu Lys Gly Leu Glu Gln Asp Pro Gln Glu Pro
260 265 270
His Gly Ala Val Arg Tyr Lys Gly Ala Leu Ser Thr Glu Glu Leu Asn
275 280 285
Phe Val Asn Gln Glu Gly Gln Ile Cys Ala Ile Pro Ala Ser Ile Gly
290 295 300
Glu Met Asp Tyr Gly Asp Phe Ile Leu Asn Gln Gln Gln Val Ile Asp
305 310 315 320
Phe Cys Lys Lys
<210> 36
<211> 336
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 36
Met Pro Ile Asn Asp Leu Lys Lys Lys Phe Glu Ile Ser Pro Gln Ala
1 5 10 15
Ala Gln Ala Ile Gly Ala Pro Ala Arg Ser Asn Ser Ser Lys Gln Ala
20 25 30
Glu His Gln Thr Glu His Leu Glu Leu Asp Thr Ser Lys Asn Arg Arg
35 40 45
Asp Arg Lys Asp Leu Asn Ala Gln Ala Thr Pro Asn Gln Gln His Thr
50 55 60
Lys Lys Leu Glu Thr Glu Val Asn Asn Gly Gly Asn Lys Ser Lys Ala
65 70 75 80
Gln Ala His Thr Pro Asp Leu Val Met Lys Lys Glu Ser Ser Val Thr
85 90 95
Pro Asn Thr Arg Lys Ser Pro Asn Glu Lys Ile Lys Ala Glu Asp Ile
100 105 110
Phe His Arg Tyr Lys Asp Arg Phe Ser Pro Ser Asp Arg Glu Leu Pro
115 120 125
Phe Glu Ile Met Asn Glu Ile Thr Asn Asn Gly Ile Ala Phe Ser Ser
130 135 140
Glu Lys Ala Pro Glu Ser His Leu Asp Lys Val Lys Asp Lys Lys Phe
145 150 155 160
Thr Leu Arg His Tyr Thr Ser Gly Asn Gly Gln Glu Lys Pro Thr Phe
165 170 175
Asn Glu Ile Gly Ser Asn Phe Asn Leu Val Asn Glu Gly Ile Lys Thr
180 185 190
Leu Lys Arg Thr Gln Gly Ser Asn Thr Asn Glu Asp Asp Trp Asn Arg
195 200 205
Leu Gly Asn Thr Ala Phe Thr Phe Phe Leu Leu Ala Ile Asp Gly Glu
210 215 220
Val Ser Asp Arg Lys Phe Leu Ser Asn Thr Thr His Phe Ala Glu Ile
225 230 235 240
Asp Ile Glu Asn Pro Ala Glu Leu Lys Glu Leu Gly Leu Asp Glu Thr
245 250 255
Glu Phe Phe Ala Ser Pro Asp Leu Leu His Glu Lys Asn Leu Ser Gln
260 265 270
Ala Pro Ala Val Lys Gly Lys Leu Ser Asp Leu Lys Ser Leu Leu Leu
275 280 285
Lys Gln Ser Gly Ile Lys Pro Val Gln Leu Gln Ser Leu Gly Ala Lys
290 295 300
Gly Ile Leu Glu Arg Ile Asp Ser Lys Phe Asn Gly Ser Leu Glu Ile
305 310 315 320
Lys Ile Pro Gly Asn Val Lys Val Lys Glu Trp Lys Lys Val Glu Lys
325 330 335
<210> 37
<211> 315
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 37
Met Pro Asn Ser Lys Tyr Ser Glu Lys Val Asn His Ser Ala Asn Gly
1 5 10 15
Ala Glu Lys Cys Ser Ile His Ser Asn Gln Tyr Asn Ile Asn Asn Cys
20 25 30
Thr Leu Gly Leu Gly Leu Asp Leu Asn Lys Lys Leu Arg Thr Gly Asn
35 40 45
Glu Arg Asn Ile Glu Gly Ala Gln Pro Phe Ile Pro Phe Pro Ser Lys
50 55 60
Gln Lys Gln Tyr Ser Thr Ser Pro Ile Ala Met Ala Asp Ile Leu Asn
65 70 75 80
Glu Ser Ala Leu Thr Ser Gln Pro Ile Ile Thr Asp Leu Ile Asn Pro
85 90 95
Gln Lys Ile Lys Met Ser Asp Gly Val Lys Asn Ile Leu Asn Asn Lys
100 105 110
Glu Gly Gly Gly Asp Leu Val Phe Lys Ala Leu Gln Ile Lys Pro Ser
115 120 125
Asp Glu Thr Leu Pro Phe Asn Ala Leu Lys Ile Val Asp Thr Tyr Gln
130 135 140
Glu Glu Met Pro Asn Lys Asp Met Ser Ile Ser Ala Tyr Trp Ala Pro
145 150 155 160
Gln Gly Gly Tyr Val Asp Ile Pro Ala Gln Pro Asp Ile Ser Arg His
165 170 175
Pro Gln Tyr Val Phe Thr Pro Asn Phe Ser Gly Cys Ser Phe Val Val
180 185 190
Asp Lys Met Asn Glu Asp Thr Leu Arg Val Arg His Val Gln Gly Gly
195 200 205
Gln Glu Asp Val Glu Tyr Asn Asn Gln Asn Ile Asp His Gly Met Gly
210 215 220
Met Ile Thr Ala Met Glu Phe Arg Asp Tyr Gly Tyr His Glu Ala Asp
225 230 235 240
Asp Lys Val Ile Glu Asn Thr Tyr Gly Phe Ala Phe Leu Lys Phe Asn
245 250 255
Gln Glu Lys Lys Gln Trp Gln Leu His Tyr Gln Lys Ile Ala Ala Ala
260 265 270
Pro Asn Ile Ile Asn Ile Lys Thr Lys Ser Ser Trp Leu Pro Phe Ser
275 280 285
Lys Pro Ser Ile Glu Ala Asp Thr Phe Thr Phe Lys Asn Met Lys Val
290 295 300
Pro Gly Tyr Ser Arg Lys Asn Ile Asn Asn Asn
305 310 315
<210> 38
<211> 309
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 38
Met Pro Lys Leu Thr Glu Leu Leu Ser Arg Phe Glu Asn Pro Ile Gln
1 5 10 15
Asn Gln Pro Asn His Ile Ser Lys Lys Asn Pro Ile Ser Asn Ser Lys
20 25 30
Val Leu Asn Asn Ser Glu Glu Lys Thr Ala Pro Leu Glu Leu Lys His
35 40 45
Asp Asp Ser Lys Ile Lys Ser Gln Val Ser Ile Pro Asn Leu Val Lys
50 55 60
Lys Asn Glu Lys Pro Ala Ala Ser Asn Thr Pro Asn Asn Ser His Glu
65 70 75 80
Lys Val Lys Ala Glu Asp Ile Phe Asn Arg Phe Lys Ser Lys Phe Asp
85 90 95
Pro Tyr Asp Arg Glu Leu Pro Phe Asp Ile Met Asn Lys Ile Thr Asn
100 105 110
Asn Glu Ile Lys Phe Ser Ser Glu Lys Ser Lys Asp Asp Tyr Leu Ala
115 120 125
Lys Val Lys Asp Lys Lys Phe Thr Leu Arg His Tyr Thr Ala Gly Thr
130 135 140
Gly Gln Glu Lys Pro Thr Phe Asp Glu Ile Ser Ser Asn Phe Asn Leu
145 150 155 160
Val Asn Lys Gly Ile Lys Thr Leu Asn Arg Thr Gln Gly Ser Asn Thr
165 170 175
Asn Glu Asp Asp Trp Asn Arg Leu Gly Asn Thr Ala Phe Thr Phe Tyr
180 185 190
Leu Leu Ala Ile Asp Gly Glu Val Ser Asn Arg Lys Phe Leu Ser Asn
195 200 205
Thr Thr His Phe Ala Glu Ile Asn Ile Glu Asp Ser Glu Glu Leu Lys
210 215 220
Glu Leu Gly Leu Asp Gln Ala Glu Phe Phe Ala Ser Pro Asp Leu Leu
225 230 235 240
His Glu Lys Asn Leu Ser Gln Ala Pro Ala Val Lys Gly Lys Leu Ser
245 250 255
Asp Leu Lys Ser Leu Leu Leu Lys Arg Ser Gly Ile Ser Ser Val Gln
260 265 270
Leu Gly Arg Leu Asp Ala Lys Ala Ile Leu Lys Ser Ile Asp Asn Glu
275 280 285
Phe Gly Asn Ser Leu Glu Ile Lys Ile Pro Gly Asn Val Lys Val Asn
290 295 300
Lys Trp Asn Lys Ile
305
<210> 39
<211> 340
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 39
Met Pro Arg Tyr Ser Asn Ser Gln Arg Thr Pro Thr Gln Ser Thr Lys
1 5 10 15
Asn Thr Arg Arg Thr Ser Pro Ser Ser Asn Ser Ser Thr Glu His Leu
20 25 30
Ser Leu Ser Asn Ala Pro Thr Asn Asp Ser Ser Val Arg Gln Glu Val
35 40 45
Lys Glu Lys Phe Ile Trp Glu Gly His Trp Glu Gly His Met Glu Ala
50 55 60
Ile Glu Lys Ala Ser Ile Leu Gly Asn Phe Ala Val Ser Phe Arg Ala
65 70 75 80
Ala Gly Lys Pro Thr Leu Glu Ala Leu Gly Lys Gly Ala Ala Ala Lys
85 90 95
Gly His Asp Ile Leu Glu Lys Thr Ile Lys Pro Gly Ser Ile Glu Lys
100 105 110
Ala Tyr Pro Glu Asn Glu Ala Ser Asp Val Ile Lys Lys Val Arg Glu
115 120 125
Ala Gly Ile Glu Gly Tyr Val Gly His Trp Asn Lys Glu Thr Gly Arg
130 135 140
Leu Glu Gly Ile Tyr Met Ser Ser Gly His Gly Leu Pro Asn Gly Gln
145 150 155 160
Val Asn Gly Lys Ile Tyr Pro Ile Asp Leu Asn Asn Leu Glu Ala Ser
165 170 175
Leu Ala Pro Leu Lys Glu Lys Lys Asn Trp Ala Ala Leu Pro Phe Thr
180 185 190
Gly Asp Tyr Asp Met His Asp Met Ile Ser Phe Thr Thr Gln Pro His
195 200 205
Ser Val Pro Ser Asn Ser Ser Glu Glu Lys Lys Ile Ile Asp Arg Ile
210 215 220
Asn Glu Tyr Ile Ala Lys Ser Asp Ser Asn Arg Pro Phe Glu Asp Ile
225 230 235 240
Glu His Asn Val Ile Arg His Gly Pro Gln Val Ser Tyr Pro Ala Phe
245 250 255
Ala Met Asp Lys Glu Lys Lys Glu Ile Lys Glu Arg Gly Gly Ile Val
260 265 270
Lys Ala Val Ala Glu Pro Gly Glu Phe Pro Val Ala Ile Val Ser Lys
275 280 285
Gly Lys Trp Thr Ile Ala Asn Asn Ile Asn Glu Leu Glu Gln Phe Tyr
290 295 300
Asn Ser Ile Gly Ala Lys Met Lys Ala Ser Trp Lys Pro Gly Ala Gly
305 310 315 320
Asn Pro Gly Phe Val Ser Asn Pro Gln Lys Pro Gly Met Ala Arg Phe
325 330 335
Ser Arg Lys Lys
340
<210> 40
<211> 280
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 40
Met Phe Ser Thr Tyr Ser Ser Lys Asn Asp Asn Gln Thr Ile Asn Lys
1 5 10 15
Ile Asn Thr Glu Glu Lys His Glu Asn Thr Glu Thr Asp Asn His Leu
20 25 30
Glu Ile Asn Leu Glu His Thr Gly Lys Ser Lys Pro Asp Ile Glu Pro
35 40 45
Lys Asp Val Thr Thr Gly Thr Ile Asn Ala Gly Thr Leu Leu Tyr Lys
50 55 60
Thr Thr Ala Ile Pro Glu Phe Leu Asp Asn Ala Lys Ser Leu Gly Leu
65 70 75 80
Ala Glu Tyr Glu Lys Arg His Lys Asp Ile Gln Asp Tyr Leu Asn Leu
85 90 95
Gly Lys Ala Glu Asp Ala Glu Lys Leu Lys Asn Lys Ser Gln Trp Ala
100 105 110
Gly Gln Tyr Phe Ala Leu Glu Lys Ser Tyr Asp Glu Tyr Ala Asn Glu
115 120 125
Ala Pro Asp Ser Tyr Asn Asn Leu Leu Lys Asn Ala Gly Lys Asp Leu
130 135 140
Leu Glu Asn Thr Glu Glu Val Lys Val Phe Leu Tyr Thr Phe Lys Val
145 150 155 160
Thr Lys Asp Ile Lys Val Leu Lys Pro His Asn Asn Ser Asn Ser Tyr
165 170 175
Tyr Val Gly Asp Thr Glu Gly Trp Glu Lys Ala Lys Glu Ile Met Asn
180 185 190
Asp Val Gln Ser Gln Ser Glu Lys Asn Asp Asn Pro Phe Pro Glu Leu
195 200 205
Lys Asn Leu Glu Asp Lys Asn Phe Leu Leu Glu Glu Leu Gly Glu Lys
210 215 220
Gly Tyr Ala Trp Met Gly Pro Leu His Ala Lys Glu Gly Ala Glu Lys
225 230 235 240
Gly Thr Glu Phe Ser Tyr Glu Leu Ala Ile Ser Pro Asn Leu Leu Arg
245 250 255
Gln His Leu Thr Leu Glu Ser Glu Glu Leu Leu Gly Thr Tyr Lys Asn
260 265 270
Arg Tyr Gly Tyr Trp Asp Lys Lys
275 280
<210> 41
<211> 138
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 41
Met Lys Lys Thr Asp Glu Lys Tyr Gly Gln Tyr Glu Tyr Lys Asp Glu
1 5 10 15
Asp Ile Thr Ser Tyr Pro Ile Ala Trp Thr Asn Pro Asp Asn Gly Lys
20 25 30
Ile Tyr Ile Gly Ile Asn Ser Pro Glu Tyr Ser His Leu Asn Asn Lys
35 40 45
Gly Glu Ser Glu Leu Asn Leu Ala Lys Ile Ile Ser Thr Ile Ile His
50 55 60
Glu Ser Leu His Ala Ser Ser His Gln His Lys Gly Leu Gln Ser Gln
65 70 75 80
Thr Asp Thr Gly Ala Asp Asn Leu Asn Tyr Asp Glu Tyr Val Thr Asp
85 90 95
Tyr Phe Ala Arg Glu Val Tyr Lys Gln Ile Leu Pro Asp Lys Asp Tyr
100 105 110
Val Ala Asn Cys Phe Thr Lys Gly Leu Gly Gly Glu Asn Lys Ile Trp
115 120 125
Gly Gly Asn Ile Val Glu Phe Met Ile Gln
130 135
<210> 42
<211> 539
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 42
Met Val Tyr Glu Tyr Asp Lys Thr Ile Glu Arg Arg Arg Asn Pro Ser
1 5 10 15
Ile Gln Leu Asn Asn Asn Glu Lys Ser Ser Glu Gln Ala Leu Glu Leu
20 25 30
Ser Gln Asn Asn Pro Leu Leu His Asp Leu Ile Thr Ser Asn Asn Leu
35 40 45
Arg Lys Glu Ala Ala Val Phe Ala Lys Arg Ile Gly Pro Ser Tyr Gln
50 55 60
Glu Ile Leu Asp Glu Leu Glu His Leu His His Leu Ser Gly Asn Glu
65 70 75 80
Gln Leu Ala Ala Gly Phe Glu Leu His Arg Arg Ile Thr His Tyr Leu
85 90 95
Glu Glu His Pro Asp Ser Lys Arg Asn Thr Ala Leu Arg Arg Thr Gln
100 105 110
Thr Gln Phe Gly Asp Leu Met Phe Thr Gly Thr Leu Gln Lys Ile Arg
115 120 125
His Ser Leu Leu Glu Met Ala Glu Thr Arg Pro Glu Met Ala Ser His
130 135 140
Ile Tyr Gln Ile Ala Arg Glu Glu Val Lys Gly Asn Thr Pro Gly Leu
145 150 155 160
Thr Asp Leu Met Val Arg Trp Val Lys Glu Asp Pro Tyr Leu Ala Ala
165 170 175
Lys Thr Gly Tyr Gln Gly Lys Ile Pro Asn Asp Leu Pro Phe Glu Pro
180 185 190
Lys Phe His Val Glu Leu Gly Ala Gln Phe Asp Asp Phe Lys Lys Trp
195 200 205
Leu Asp Thr Ala Gln Ser Lys Glu Leu Leu Thr His Thr Arg Leu Asp
210 215 220
Glu Gln Asn Lys Gln Val His Leu Gly Tyr Ser Tyr Asn Glu Leu Leu
225 230 235 240
Asp Met Thr Gly Val Glu Ser Val Gln Met Ala Val Tyr Phe Leu Lys
245 250 255
Glu Ala Ala Lys Gln Ala Glu Pro Gly Ser Thr Lys Ser Gln Glu Asp
260 265 270
Ile Leu Leu His Arg Phe Ala Asn Pro Thr Tyr Leu Ala Gln Leu Glu
275 280 285
His Ser Arg Leu Ala Gln Ile Glu Ala Ile Tyr His Ser Ser His Asp
290 295 300
Thr Asp Val Thr Ala Trp Asp Gln Gln Phe Ala Ser Asp Ala Leu Thr
305 310 315 320
Gln Phe Asn His Gln Leu Asn Asn Thr Val Asp Leu Asn Ser Gln Leu
325 330 335
Ser Leu Leu Leu Lys Asp Arg Gln Gly Leu Leu Ile Gly Glu Ser His
340 345 350
Gly Ser Asp Leu Asn Gly Leu Arg Phe Val Glu Glu Gln Met Glu Val
355 360 365
Leu Lys Ala His Gly Val Thr Val Ile Gly Leu Glu His Leu Arg Ser
370 375 380
Asp Leu Ala Gln Pro Leu Ile Asp Lys Phe Leu Ala Ser Gly Asn Glu
385 390 395 400
Pro Met Pro Ala Glu Leu Ala Ala Leu Leu Lys Thr Lys His Leu Ser
405 410 415
Ala Asn Leu Phe Glu Gln Ala Arg Ser Lys Gln Met Lys Ile Ile Ala
420 425 430
Leu Asp Asn Asn Ser Thr Thr Arg Pro Thr Val Glu Gly Thr Gln His
435 440 445
Gly Leu Met Tyr Arg Ala Gly Ala Ala Asn Asn Val Ala Val Glu Arg
450 455 460
Leu Arg Gln Leu Pro Ala Gly Glu Lys Phe Val Ala Ile Tyr Gly Asn
465 470 475 480
Ala His Leu Gln Ser His Glu Gly Ile Asp His Phe Leu Pro Gly Ile
485 490 495
Thr His Arg Leu Gly Leu Pro Ala Leu Lys Val Asp Glu Asn Asn Arg
500 505 510
Phe Thr Ala Gln Val Asp Asn Ile Asn Gln Arg Lys Arg Tyr Asp Asp
515 520 525
Val Val Glu Leu Pro Arg Ile Gln Leu Thr Ser
530 535
<210> 43
<211> 323
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 43
Met Glu His Glu Tyr Ser Glu Lys Glu Lys Pro Gln Lys Cys Pro Ile
1 5 10 15
Gln Leu Arg Asp Ser Ile Glu His Asp Lys Glu Asp Ile Asn Thr Thr
20 25 30
Thr Pro Leu Glu Leu Asn Ser Gln Tyr Thr Asn Arg Lys Arg Ala Gly
35 40 45
Leu Arg Glu Arg Phe Ser Thr Thr Leu Gln Arg Asn Leu Pro Gly His
50 55 60
Ser Met Leu Asp Arg Glu Leu Thr Thr Asp Gly Met Lys Asn Gln Glu
65 70 75 80
Ser Arg Phe Ser Pro Ala Met Ile Met Asp Arg Met Met His Phe Gly
85 90 95
Val Arg Thr Arg Leu Gly Lys Val Arg Asn Ser Ala Ser Lys His Gly
100 105 110
Gly Gln Val Thr Phe Lys Phe Ala Gln Thr Lys Gly Thr Phe Leu Asp
115 120 125
Gln Ile Met Lys His Lys Asp Thr Ser Gly Gly Val Cys Glu Ser Ile
130 135 140
Ser Ala His Trp Ile Ser Ala His Ala Lys Gly Glu Ser Ile Phe Asp
145 150 155 160
Gln Leu Tyr Val Gly Gly Gln Lys Gly Lys Phe His Ile Asp Ser Leu
165 170 175
Val Ser Ile Lys Gln Leu Gln Met Asp Ser Tyr Leu Asp Asp Glu Gln
180 185 190
Ser Thr Met Thr Glu Tyr Trp Leu Gly Thr Gln Gly Ile Gln Pro Ile
195 200 205
Met Gln Lys Asn Asp Val Asp Glu His Ser Ser Lys Val Val Gly Gln
210 215 220
Thr Gly Asn Lys Gly Thr Thr Asp Leu Leu Arg Ala Ile Leu Asp Thr
225 230 235 240
Gly Asp Lys Gly Ser Gly Tyr Lys Lys Ile Ser Phe Leu Gly Lys Met
245 250 255
Ala Gly His Thr Val Ala Ala Tyr Val Asp Asp Gln Lys Gly Val Ile
260 265 270
Phe Phe Asp Pro Asn Phe Gly Glu Phe Ser Phe Pro Ser Ile Thr Ser
275 280 285
Phe Ser Arg Trp Phe Thr Asp Asp Phe Trp Pro Lys Ser Trp Tyr Asn
290 295 300
Leu Glu Ile Gly Leu Gly Gln Gln Phe Glu Val Phe Asn Tyr Glu Leu
305 310 315 320
Lys Lys Ser
<210> 44
<211> 381
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 44
Met Tyr Asp Ser Lys Lys Lys Asn Ser Glu Pro Thr Thr Lys Lys Lys
1 5 10 15
Phe Glu Arg Ser Asn Tyr Ser Gln Trp Asp Asp Ser Ile Asn His Tyr
20 25 30
Glu Asp Met Asn Arg Ala Arg Ile Lys Asn Arg Asn Asp Ile Leu Thr
35 40 45
Thr Val Asp Tyr Phe Gly Glu Lys Lys Lys Thr Met His Thr Phe Glu
50 55 60
Tyr Gln Ser Asp Ile Lys His Asp Thr Asn Phe Asn Asn Lys Asn Lys
65 70 75 80
Ser Leu Phe Glu Ser Phe Ala Ala Ser Phe Val Leu Gln Asn Pro Ser
85 90 95
Phe Phe Ser Gly Val Ile Asp Lys Leu Ser Lys Lys Leu Phe Asn Ile
100 105 110
Ile Ser Lys Ile Asp Glu Arg Asn Asn Phe Gln Lys Lys Leu Tyr Asp
115 120 125
Phe Ile Glu Lys Asp Thr Ser Pro Glu Gly Gln Phe Gly Arg Phe Thr
130 135 140
Leu Gly Lys Asn Glu Ile Leu Asn Val Leu Gln Val Lys Ser Asp Thr
145 150 155 160
Pro Gln Leu Phe Val Lys Lys Met Leu Leu Ile Lys Ser Leu Gly Ala
165 170 175
Phe Ile Ile Asp Phe Ser Ser Lys Asp Ile Gly Asn Tyr Asp Phe Ile
180 185 190
Phe Asp Gly Lys Gly Arg Glu Val Asn Asp Ile Ile Glu Lys Asn Arg
195 200 205
Pro Thr Asn Leu Phe Lys Val Arg Gly Arg Thr Asn Ile Lys Ser Ser
210 215 220
Gln His Arg Ser Asp Ile Gly Ile Leu Asp Thr Pro Thr Phe Asp Ser
225 230 235 240
Leu Thr Glu Glu Gln Lys Ser Phe Leu Thr Ile Pro Glu Leu Thr Lys
245 250 255
Arg Arg Pro Leu Phe Arg Thr Phe Thr His Glu Leu Asp Ala Glu Asp
260 265 270
Lys Arg Val Val Glu Ser Val Phe Val Asn Arg Thr Phe Asp Cys Asp
275 280 285
Ser Pro Leu Ile Gly Ser Val Ser Gly Ser Thr Ser Cys Val Leu Val
290 295 300
Ala Ala Asp Ile Leu Phe Pro Asp Met Thr Met Val Glu Arg Lys Lys
305 310 315 320
Leu Ala Ile Ala Thr Phe Ala Phe Leu Val Gly Gly Gly Tyr His Ser
325 330 335
Ala Thr Glu Val Phe Asp Val Ala Tyr Pro Gly Leu Asp Leu Asn Lys
340 345 350
Glu Ile Glu Glu Leu Ile Glu Asn Asn Pro Ile Gln Glu Asn Ala Gly
355 360 365
Val Ala Thr Leu Arg Gln Leu Ile Gly Asn Ser Gly Phe
370 375 380
<210> 45
<211> 308
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 45
Met Pro Ile Ser Asn Leu Ala Lys Glu Ser Glu Val Arg Ala Val Lys
1 5 10 15
Asp Ile Pro Cys Lys Asn Ile Glu Thr Asp Asn His Leu Glu Ile Gly
20 25 30
Leu Ser Ser Gly Leu Ser Arg Ser Lys Asp Thr Ser Lys Phe Lys Lys
35 40 45
Asn Ser Ile Asn Thr Ile Lys Leu Ile Asp Asp Ile Ile Ala Leu His
50 55 60
Asn Asp Pro Lys Gly Asn Lys Leu Leu Trp Asn Asp Asn Trp Gln Asp
65 70 75 80
Lys Ile Ile Asn Arg Asp Leu Ala Asn Ile Phe Glu Lys Ile Asp Glu
85 90 95
Ser Val Ser Glu Leu Gly Gly Leu Glu Met Tyr Gln Glu Met Val Gly
100 105 110
Val Asn Pro Tyr Asp Pro Thr Glu Pro Val Cys Gly Leu Ser Ala Gln
115 120 125
Asn Ile Phe Lys Leu Met Thr Glu Gly Glu His Ala Val Asp Pro Val
130 135 140
Glu Met Ala Gln Thr Gly Lys Ile Asp Gly Asn Glu Phe Ala Glu Ser
145 150 155 160
Val Asp Gln Leu Ser Ser Ala Lys Asn Tyr Val Ala Leu Val Asn Asp
165 170 175
Arg Arg Leu Gly His Met Phe Leu Ile Asp Ile Pro Ser Asn Asp Gln
180 185 190
Glu Thr Val Gly Tyr Ile Tyr Gln Ser Asp Leu Gly Gln Gly Ala Leu
195 200 205
Pro Pro Leu Lys Ile Ala Asp Trp Leu Asn Ser Arg Gly Lys Asp Ala
210 215 220
Val Ser Leu Asn Lys Leu Lys Lys Leu Leu Ser Arg Glu Phe Asn Leu
225 230 235 240
Leu Ser Asp Asp Glu Lys Arg Ala Leu Ile Ser Glu Thr Leu Asp Ile
245 250 255
His Lys Asp Val Ser Asn Val Glu Leu Asp Arg Ile Lys Arg Asp Arg
260 265 270
Gly Val Asp Ile Tyr Leu Thr Glu Tyr Asp Val Asn Asn Phe Tyr Glu
275 280 285
Asn Ile Glu Thr Leu Lys Ser Lys Leu Ser Asn Tyr Asp Lys Lys Leu
290 295 300
Ser Lys Pro Lys
305
<210> 46
<211> 295
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 46
Met Leu Ala Asn Val Leu Pro Asn Leu Ala Ser Phe Leu Lys Tyr Glu
1 5 10 15
Lys Glu Thr Pro Leu Phe Phe Ile Glu Asp Gly Phe Asn Phe Gln Asn
20 25 30
Leu Asn Pro Gly Arg Val Pro Leu Ile Lys Thr Pro Glu Gln Arg Lys
35 40 45
Ala Gly Asp Thr Gln Ser Pro Ala Phe Leu Cys Ser Gly Val Ile Leu
50 55 60
Arg Gly Thr Ile His Ser Asn Asp Tyr Lys Phe Trp Gln Pro Ser Pro
65 70 75 80
Ser Ser Ile Lys Ser Gly Gly Val Ser Phe Ser Tyr Leu Arg Lys Asp
85 90 95
Ala Lys Phe Lys Arg Leu Ala Tyr Gly Tyr Lys Asn Gly Phe Ile Ile
100 105 110
Phe Pro Glu His Ile Ala Pro Glu Asp Arg Val Asp Phe Ser Val Leu
115 120 125
Cys Ala Phe Pro Ile Asp Gly Tyr Thr Asn Glu Arg Ala Asn Gln Gly
130 135 140
Cys Gly Glu Asn Ile Thr Lys Ala Lys Asp Lys Gly Lys Ser Cys Gln
145 150 155 160
Glu Gln Asn Val Thr Asn Ser Asp Asp Trp Ile Lys Asn Tyr Arg Lys
165 170 175
Val Asn Ser Gln Asp Phe Phe Gln Cys Gly Phe Asn Val Thr Lys Asp
180 185 190
Val Asn Asn Pro Ala Ile Ala Phe Tyr Gln Met Leu Glu Ser Ile Lys
195 200 205
Lys Leu Pro Arg Thr Pro Asn Thr Pro Pro Lys Gln Asn Glu Ile Arg
210 215 220
Ile Ser Thr Trp Glu Glu Ser Asp Pro Asn Lys Leu Pro Ile Glu Ala
225 230 235 240
Leu Phe Tyr Ser Glu Asn Ser Gly Leu Ala Asp Ala Gln Lys Asp Gln
245 250 255
Arg Asp Tyr Lys Asn Ala Thr Gly Lys Phe Leu Pro Ile Val Lys Met
260 265 270
Leu Leu Pro Arg Thr Leu Asn Glu Asp Ala Leu Phe Lys Phe Asn Ile
275 280 285
Lys Asp Gln Val Ile Asn Pro
290 295
<210> 47
<211> 50
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 47
Met Met Arg Glu Tyr Ser Asn Glu Asp Asp Phe Ile Lys Glu Lys Thr
1 5 10 15
Asn Leu Val Lys Ser Glu Asn Val Glu Ala Asp Asn Tyr Leu Glu Thr
20 25 30
Glu Tyr Leu Thr Tyr Leu Ala Lys Leu Ile Gly Met Thr Glu Arg Glu
35 40 45
Asn His
50
<210> 48
<211> 50
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 48
Met Phe Gln Asn Arg Ile Arg Asn Glu Lys Thr Thr Gln Ser Gly Lys
1 5 10 15
Gly Lys Thr Leu Asp Arg Met Thr Asp Ser Leu Tyr Leu Glu Ile Pro
20 25 30
Asn Val Glu Ala Val Thr Leu Ala Tyr Gln Lys Leu Thr Ser Lys Tyr
35 40 45
Arg Lys
50
<210> 49
<211> 50
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 49
Met Glu Arg Glu Tyr Ser Glu Lys Gln Lys Asn Pro Ser Lys Leu Ser
1 5 10 15
Arg Lys Thr Ala Ile Ser Glu Arg Ile Ala Ala Leu Glu Arg Ser Gly
20 25 30
Leu Ser Asn Ser Asn Gln Pro Val Pro Gln Phe Ala Arg Pro Tyr Thr
35 40 45
Ser Asn
50
<210> 50
<211> 50
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 50
Met Ser Asn Tyr Glu Tyr Asp Ile Val Thr Gln His Asp Thr Tyr Gln
1 5 10 15
Ile Lys Asp Asn Glu Tyr Thr Val Val Asn Gly Lys Tyr Trp Gln Tyr
20 25 30
Glu Gln Glu Gly Asn Lys Asn Asn Asn Lys Val Ser Ile Ser Leu Met
35 40 45
Lys Glu
50
<210> 51
<211> 50
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 51
Met Glu His Glu Tyr Asn Glu Lys Glu Lys Gln Arg Asn Ser Ala Ile
1 5 10 15
Lys Leu Asn Asp Ala Ile Arg Asn Asn Glu Glu Asn Met Asp Met Thr
20 25 30
Ser Pro Leu Glu Leu Asn Phe Gln Asn Thr Asn Arg Lys Ser Arg Gly
35 40 45
Leu Arg
50
<210> 52
<211> 50
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 52
Met Pro Asn Lys Lys Tyr Ser Glu Asn Thr His Gln Gly Lys Lys Pro
1 5 10 15
Leu Met Lys Ser Glu Ala Asn Asn Glu His Asp Ile Gln Asn Ser Ser
20 25 30
Leu Gly Ile Gly Leu Asp Leu Asn Ser Met Met Gly Asn Ser Ser Thr
35 40 45
Ser Leu
50
<210> 53
<211> 50
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 53
Met Glu Arg Glu Tyr Ser Glu Lys Glu Lys His Lys Lys Arg Pro Ile
1 5 10 15
Gln Leu Arg Asn Ser Ile Glu Gln His Glu Glu Glu Thr Ala Asn Asn
20 25 30
Ser Leu Gly Leu Gly Leu Asp Leu Asn Gln Ala Thr Asn Pro Pro Lys
35 40 45
Val Pro
50
<210> 54
<211> 50
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 54
Met Met Glu His Glu Tyr Ser Lys Glu Glu Glu Lys Lys Arg Gln Gln
1 5 10 15
Ser Lys Pro Asn Asn Ala Thr His Asp Glu Ser Asn Leu Pro Leu Glu
20 25 30
Leu Glu Lys His Phe Asn Ala Arg Thr Pro Ala Thr Ala His Ser Lys
35 40 45
Trp Phe
50
<210> 55
<211> 50
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 55
Met Leu Lys Tyr Ala Asn Pro Gln Ala Val Pro Thr Gln Arg Thr Lys
1 5 10 15
Asn Thr Ala Lys Lys Pro Ser Ser Ser Ser Ser Phe Asp Gly Gln Leu
20 25 30
Glu Leu Ser Asn Gly Glu Trp Ser Lys His Ser Glu Met Gly Leu Lys
35 40 45
Arg Gly
50
<210> 56
<211> 50
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 56
Met Met Arg Glu Tyr Ser Asn Glu Asp Asp Cys Thr Lys Glu Lys Thr
1 5 10 15
Asn Leu Val Lys Ser Glu Asn Val Glu Ala Asp Asn Tyr Leu Glu Met
20 25 30
Glu His Leu Thr Tyr Leu Ala Lys Leu Ile Ser Met Thr Glu Arg Glu
35 40 45
Asn His
50
<210> 57
<211> 50
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 57
Met Ile Phe Lys Met Leu Asn Leu Ala Val Phe Tyr Leu Leu Gly Asn
1 5 10 15
Ile Phe His Tyr Leu Ile Cys Gln Lys Phe Ile Cys Tyr Phe Cys Ser
20 25 30
Val Leu Lys Ser Val Thr Met Phe Leu Thr Lys Val Ala Val Gln Ile
35 40 45
Ala Leu
50
<210> 58
<211> 50
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 58
Met Glu Arg Glu Tyr Ser Glu Lys Pro Lys Asn Leu Ser Gln Leu Ser
1 5 10 15
Arg Lys Thr Ala Ile Ser Glu Arg Arg Ala Met Phe Glu Arg Asn Ala
20 25 30
Ser Ser Asn Asn Glu Gln Pro Val Pro Gln Phe Ala Arg Ser Tyr Thr
35 40 45
Ser Asn
50
<210> 59
<211> 50
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 59
Met Lys Tyr Asp Pro Arg Leu Arg Thr Trp Val Glu Asp Asp Phe Asp
1 5 10 15
Tyr Glu Lys Asn Phe Lys Lys Gln Thr Asp Tyr Ile Asn Tyr Lys Asp
20 25 30
Leu Glu Lys Gln Leu Lys Glu Asn Val Asp Tyr Tyr Ala Leu Leu Asp
35 40 45
Glu Asn
50
<210> 60
<211> 50
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 60
Met Pro Asn Lys Lys His Ser Glu Asn Thr His Gln Gly Arg Lys Pro
1 5 10 15
Leu Ile Lys Ser Glu Ala Asn Asn Glu His Asp Ile Glu Asn Ser Ser
20 25 30
Leu Gly Ile Gly Leu Asp Leu Asn Ser Thr Ile Gly Asn Asn Ser Ala
35 40 45
Ser Leu
50
<210> 61
<211> 50
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 61
Met Glu Arg Glu Tyr Ser Glu Lys Glu Lys His Lys Lys Arg Pro Ile
1 5 10 15
Gln Leu Arg Asn Ser Ile Glu Gln His Glu Glu Glu Thr Ala Asn Asn
20 25 30
Ser Leu Gly Leu Gly Leu Asp Leu Asn Gln Ala Thr Asn Pro Pro Lys
35 40 45
Val Pro
50
<210> 62
<211> 50
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 62
Met Met Glu His Glu Tyr Ser Lys Glu Glu Glu Lys Lys Arg Gln Gln
1 5 10 15
Ser Lys Pro Asn Asn Ala Thr His Asp Glu Ser Asn Leu Pro Leu Glu
20 25 30
Leu Glu Lys His Ser Asn Ala Arg Thr Ser Ala Thr Ala Tyr Ser Lys
35 40 45
Trp Phe
50
<210> 63
<211> 50
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 63
Met Ser Asn Tyr Glu Tyr Asp Ile Val Thr Gln His Asp Thr Tyr Gln
1 5 10 15
Ile Lys Asp Asn Glu Tyr Thr Val Val Asn Gly Lys Tyr Trp Gln Tyr
20 25 30
Glu Gln Glu Gly Asn Lys Asn Asn Asn Lys Ile Ser Ile Ser Leu Met
35 40 45
Lys Asp
50
<210> 64
<211> 50
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 64
Met Glu His Glu Tyr Asn Glu Lys Glu Lys Gln Arg Asn Ser Ala Ile
1 5 10 15
Lys Leu Asn Asp Ala Ile Arg Asn Asn Glu Glu Asn Met Asp Met Thr
20 25 30
Ser Pro Leu Glu Leu Asn Ser Gln Asn Thr Asn Arg Lys Ser Arg Gly
35 40 45
Leu Arg
50
<210> 65
<211> 50
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 65
Met Phe Lys Tyr Asp Thr Ser Glu Lys Met Ala Lys Phe Gly Lys Gly
1 5 10 15
Lys Thr Ser Asp Gly Met Leu Leu Asp Thr Leu Tyr Leu Glu Ile Pro
20 25 30
Asp Glu Lys Ala Val Met Ser Ala Tyr Lys Ser Gln Ile Leu Asp Glu
35 40 45
Leu Arg
50
<210> 66
<211> 50
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 66
Met Leu Lys His Ala Asn Pro Gln Thr Val Ser Thr Gln Arg Thr Lys
1 5 10 15
Ser Thr Ala Lys Lys Pro Ser Ser Ser Ser Ser Phe Asp Arg Gln Phe
20 25 30
Glu Leu Ser Asn Ser Glu Asn Gln Pro Gly Glu Gly Asn Lys Asp Trp
35 40 45
Thr Ile
50
<210> 67
<211> 50
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 67
Met Pro Arg Tyr Ala Asn Tyr Gln Ile Asn Pro Lys Gln Asn Thr Lys
1 5 10 15
Asn Ser His Gly Lys Ser Ser Ser Ser Asn Phe Ser Ser Gly Tyr Phe
20 25 30
Ser Ser Ser Asn Asn Ser Leu Asp Asp Ser Leu Ile Arg Gln Gln Val
35 40 45
Lys Arg
50
<210> 68
<211> 50
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 68
Met Arg Glu Tyr Ser Lys Glu Asp Asp Cys Val Lys Glu Lys Thr Asn
1 5 10 15
Leu Ala Glu Ser Glu Asn Val Glu Ala Asp Asn Tyr Leu Glu Met Asp
20 25 30
Cys Leu Asn Tyr Leu Ala Lys Leu Asn Gly Met Pro Glu Arg Lys Asp
35 40 45
His Ser
50
<210> 69
<211> 50
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 69
Met Pro Ile Ile Gly His Lys Glu Asp Leu Ile Arg Thr Glu Arg Ser
1 5 10 15
Ser Val Asp Leu Thr Arg Ser Ser Asn Asn Arg Gln Thr Asp Asn Leu
20 25 30
Glu Leu Asn Ile Pro Gln His Lys Arg Asp Asn Lys Asp Ile Glu His
35 40 45
Ala Val
50
<210> 70
<211> 50
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 70
Met Ile Ser Thr Phe Asp Pro Ala Ile Cys Ala Gly Thr Pro Thr Val
1 5 10 15
Thr Val Leu Asp Asn Arg Asn Leu Thr Val Arg Glu Ile Val Phe His
20 25 30
Arg Ala Lys Ala Gly Gly Asp Thr Asp Thr Leu Ile Thr Arg His Gln
35 40 45
Tyr Asp
50
<210> 71
<211> 50
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 71
Met Glu Arg Glu Tyr Asn Lys Lys Glu Lys Gln Lys Lys Ser Ala Ile
1 5 10 15
Lys Leu Asp Asp Ala Val Gly Asn Asn Glu Glu Asn Met Asp Met Thr
20 25 30
Ser Pro Leu Glu Leu Asn Ser Gln Tyr Thr Asn Arg Lys Arg Pro Gly
35 40 45
Leu Arg
50
<210> 72
<211> 50
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 72
Met Val Tyr Glu Tyr Ala Lys Thr Asn Asp Arg Lys Arg Lys Leu Ser
1 5 10 15
Thr Gln Ser Asp Asn Tyr Glu Glu Lys Ser Phe Ser Pro Val Leu Asp
20 25 30
Leu Ser Arg Asn Asn Gln Asn Thr Pro Asn Met Glu Asp Glu Tyr Glu
35 40 45
Thr Pro
50
<210> 73
<211> 50
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 73
Met Val Phe Glu His Asp Lys Thr Val Glu Arg Lys Arg Lys Pro Ser
1 5 10 15
Ile Gln Leu Gly Asn Asp Lys Glu Lys Ser Ser Glu Gln Ala Leu Glu
20 25 30
Leu Pro Gln Ser Lys Gln Asn Asn Pro Leu Leu His Asp Leu Ile Thr
35 40 45
Ser Asn
50
<210> 74
<211> 50
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 74
Met Lys Gly Ile Glu Gly Val Ile Met Leu Ser His Asp Ile Leu Pro
1 5 10 15
Glu Lys Leu Leu Val Ser Glu Lys Lys His Glu Asn Val Gly Ser Tyr
20 25 30
Phe Ser Asp Asp Ile Gly Glu Gln Ser Glu Gln Thr Glu Val Ser His
35 40 45
Phe Asn
50
<210> 75
<211> 50
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 75
Met Pro Asn Lys Lys Tyr Ser Glu Asn Thr His Gln Gly Lys Lys Pro
1 5 10 15
Leu Ile Lys Ser Glu Ala Asn Asn Glu His Ala Ile Asp Asn Ser Pro
20 25 30
Leu Gly Ile Gly Leu Asp Leu Asn Ser Ile Leu Gly Asn Asn Ser Ala
35 40 45
Ser Leu
50
<210> 76
<211> 50
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 76
Met Glu Arg Glu Tyr Ser Glu Lys Glu Lys His Lys Lys His Pro Ile
1 5 10 15
Gln Leu Arg Asp Ala Ile Glu Gln His Ala Glu Glu Thr Ala Asn Asn
20 25 30
Ser Leu Gly Leu Gly Leu Asp Leu His Gln Ala Ile Asn Thr Pro Lys
35 40 45
Val Pro
50
<210> 77
<211> 50
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 77
Met Val His Glu Tyr Ser Ile Asn Asp Arg Gln Lys Arg His Ser Phe
1 5 10 15
Ser Ser Ala Asn Pro Ile Asp Pro Glu Val Thr Asn Arg Glu Asn Ser
20 25 30
Arg His Arg Phe Pro Lys Asp Asn Tyr Asn Lys Gly His Gly Asp Leu
35 40 45
Phe Tyr
50
<210> 78
<211> 50
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 78
Met Leu Lys Tyr Ala Asn Pro Gln Thr Val Ala Thr Gln Arg Thr Lys
1 5 10 15
Asn Thr Ala Lys Lys Pro Pro Ser Ser Thr Ser Phe Asp Gly His Leu
20 25 30
Glu Leu Ser Asn Gly Glu Asn Gln Pro Tyr Glu Gly His Lys Ile Arg
35 40 45
Lys Ile
50
<210> 79
<211> 50
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 79
Met Pro Arg Tyr Ala Asn Tyr Gln Ile Asn Pro Lys Gln Asn Ile Lys
1 5 10 15
Asn Ser His Gly Lys Ser Ser Ser Ser Asp Phe Ser Ser Gly Tyr Leu
20 25 30
Ser Phe Ser Asn Asn Ser Leu Asp Asp Pro Phe Ile Arg Gln Gln Val
35 40 45
Lys Arg
50
<210> 80
<211> 50
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 80
Met Pro Asn Lys Lys Tyr Ser Glu Asn Thr His Gln Gly Lys Asn Pro
1 5 10 15
Leu Met Lys Ser Gly Ala Asn Asn Glu His Asp Leu Gln Asp Ser Pro
20 25 30
Leu Gly Ile Gly Leu Asp Leu Asn Ser Met Leu Val Asn Ser Ser Thr
35 40 45
Ser Leu
50
<210> 81
<211> 50
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 81
Met Leu Ser Thr Glu Lys His Asn Lys Asp Thr Lys His Pro Arg Asn
1 5 10 15
Arg Glu Lys Lys Phe Ser Ile Gln Pro Glu Asn Ser Thr Gln Asp Asp
20 25 30
Glu Asp Ile Lys Asn Asn Ser Leu Gly Val Gly Leu Asp Leu Asp Gln
35 40 45
Met Ile
50
<210> 82
<211> 50
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 82
Met Pro Ile Asn Asp Leu Lys Lys Lys Phe Glu Ile Ser Pro Gln Ala
1 5 10 15
Ala Gln Ala Ile Gly Ala Pro Ala Arg Ser Asn Ser Ser Lys Gln Ala
20 25 30
Glu His Gln Thr Glu His Leu Glu Leu Asp Thr Ser Lys Asn Arg Arg
35 40 45
Asp Arg
50
<210> 83
<211> 50
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 83
Met Pro Asn Ser Lys Tyr Ser Glu Lys Val Asn His Ser Ala Asn Gly
1 5 10 15
Ala Glu Lys Cys Ser Ile His Ser Asn Gln Tyr Asn Ile Asn Asn Cys
20 25 30
Thr Leu Gly Leu Gly Leu Asp Leu Asn Lys Lys Leu Arg Thr Gly Asn
35 40 45
Glu Arg
50
<210> 84
<211> 50
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 84
Met Pro Lys Leu Thr Glu Leu Leu Ser Arg Phe Glu Asn Pro Ile Gln
1 5 10 15
Asn Gln Pro Asn His Ile Ser Lys Lys Asn Pro Ile Ser Asn Ser Lys
20 25 30
Val Leu Asn Asn Ser Glu Glu Lys Thr Ala Pro Leu Glu Leu Lys His
35 40 45
Asp Asp
50
<210> 85
<211> 50
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 85
Met Pro Arg Tyr Ser Asn Ser Gln Arg Thr Pro Thr Gln Ser Thr Lys
1 5 10 15
Asn Thr Arg Arg Thr Ser Pro Ser Ser Asn Ser Ser Thr Glu His Leu
20 25 30
Ser Leu Ser Asn Ala Pro Thr Asn Asp Ser Ser Val Arg Gln Glu Val
35 40 45
Lys Glu
50
<210> 86
<211> 50
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 86
Met Phe Ser Thr Tyr Ser Ser Lys Asn Asp Asn Gln Thr Ile Asn Lys
1 5 10 15
Ile Asn Thr Glu Glu Lys His Glu Asn Thr Glu Thr Asp Asn His Leu
20 25 30
Glu Ile Asn Leu Glu His Thr Gly Lys Ser Lys Pro Asp Ile Glu Pro
35 40 45
Lys Asp
50
<210> 87
<211> 50
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 87
Met Lys Lys Thr Asp Glu Lys Tyr Gly Gln Tyr Glu Tyr Lys Asp Glu
1 5 10 15
Asp Ile Thr Ser Tyr Pro Ile Ala Trp Thr Asn Pro Asp Asn Gly Lys
20 25 30
Ile Tyr Ile Gly Ile Asn Ser Pro Glu Tyr Ser His Leu Asn Asn Lys
35 40 45
Gly Glu
50
<210> 88
<211> 50
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 88
Met Val Tyr Glu Tyr Asp Lys Thr Ile Glu Arg Arg Arg Asn Pro Ser
1 5 10 15
Ile Gln Leu Asn Asn Asn Glu Lys Ser Ser Glu Gln Ala Leu Glu Leu
20 25 30
Ser Gln Asn Asn Pro Leu Leu His Asp Leu Ile Thr Ser Asn Asn Leu
35 40 45
Arg Lys
50
<210> 89
<211> 50
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 89
Met Glu His Glu Tyr Ser Glu Lys Glu Lys Pro Gln Lys Cys Pro Ile
1 5 10 15
Gln Leu Arg Asp Ser Ile Glu His Asp Lys Glu Asp Ile Asn Thr Thr
20 25 30
Thr Pro Leu Glu Leu Asn Ser Gln Tyr Thr Asn Arg Lys Arg Ala Gly
35 40 45
Leu Arg
50
<210> 90
<211> 50
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 90
Met Tyr Asp Ser Lys Lys Lys Asn Ser Glu Pro Thr Thr Lys Lys Lys
1 5 10 15
Phe Glu Arg Ser Asn Tyr Ser Gln Trp Asp Asp Ser Ile Asn His Tyr
20 25 30
Glu Asp Met Asn Arg Ala Arg Ile Lys Asn Arg Asn Asp Ile Leu Thr
35 40 45
Thr Val
50
<210> 91
<211> 50
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 91
Met Pro Ile Ser Asn Leu Ala Lys Glu Ser Glu Val Arg Ala Val Lys
1 5 10 15
Asp Ile Pro Cys Lys Asn Ile Glu Thr Asp Asn His Leu Glu Ile Gly
20 25 30
Leu Ser Ser Gly Leu Ser Arg Ser Lys Asp Thr Ser Lys Phe Lys Lys
35 40 45
Asn Ser
50
<210> 92
<211> 50
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 92
Met Leu Ala Asn Val Leu Pro Asn Leu Ala Ser Phe Leu Lys Tyr Glu
1 5 10 15
Lys Glu Thr Pro Leu Phe Phe Ile Glu Asp Gly Phe Asn Phe Gln Asn
20 25 30
Leu Asn Pro Gly Arg Val Pro Leu Ile Lys Thr Pro Glu Gln Arg Lys
35 40 45
Ala Gly
50
<210> 93
<211> 19592
<212> DNA
<213> 光杆状菌(Photorhabdus)
<400> 93
atgtctacaa gtacatctca aattgcggtt gaatatccta ttcctgtcta tcgctttatt 60
gtttctgtcg gagatgagaa aattccattt aatagtgttt caggattaga tattagttat 120
gacaccattg aataccgaga tggtgttggt aattggttca aaatgccggg tcagagtcag 180
agcactaata tcaccttgcg taaaggcgtt ttcccgggga aaacagaact gtttgattgg 240
attaactcta ttcagcttaa tcaggtagag aaaaaggata ttaccatcag tttaactaat 300
gatgcaggta ccgaattatt aatgacctgg aatgtttcta atgcttttcc cacttcattg 360
acttcacctt catttgatgc caccagtaat gatattgcag tacaggaaat tacgctgatg 420
gcagatcggg tgattatgca ggctgtttga agcattgata tttaatcatc tcatataagg 480
gaacttttat gacaaccgtt accagttatc ctggcgttta tattgaagaa ttaaatagcc 540
tggccttgtc agtttcaaat agcgccacag cggttcctgt ttttgctgtg gacgaacaaa 600
accaatatat tagtgaagat aatgcaatcc gtattaattc gtggatggat tatcttaatc 660
tgattggcaa ttttaataat gaagacaaat tagatgtttc tgtgcgtgct tattttgcca 720
atggaggtgg atattgttat ctcgtcaaaa caacgagttt agaaaaaatt attccaacct 780
tggatgatgt aaccttattg gttgctgcgg gcgaagatat taaaacgaca gtagatgttt 840
tatgtcagcc aggaaaaggg ttattcgcag tctttgatgg ccctgaaaca gagttgacta 900
tcaacggtgc ggaagaggca aaacaagcct ataccgccac accattcgct gcggtttatt 960
atccttggtt gaaagcggat tgggctaaca tagatattcc acccagtgca gtgatggcgg 1020
gagtttatgc atcggtggat ttatcccgtg gtgtatggaa agcgcctgcc aatgttgcgt 1080
tgaaaggggg cctggaacct aaatttttag tcacggatga attgcagggt gaatataaca 1140
ctggccgcgc tatcaatatg attcgtaatt tcagtaacac aggtactacg gtttggggtg 1200
caagaaccct ggaagataaa gacaattggc gttatgttcc agtgcgacgc ttgtttaatt 1260
ctgtggagcg ggatatcaag cgtgccatga gctttgctat gttcgagcct aataatcagc 1320
ctacttggga gcgggtacgg gcggcgatta gcaactacct ttatagcctg tggcaacagg 1380
ggggattagc tggcagcaaa gaagaagacg cttattttgt gcaaattggt aaaggtataa 1440
cgatgacaca ggagcagatt gatgcagggc aaatgattgt taaagtcggt ttggctgctg 1500
tacggcctgc ggaatttatc attctccagt ttacgcaaga tgtagaacag cgttaatcat 1560
atgattatga ggagttatca tgtctgctat tctgaaagcg cctggcgttt atattgaaga 1620
agacgcttcc ctagcgttgt ctgtcagtaa cagcgcgact gccgtgcctg tttttatcgg 1680
aaaatttact ccgacagtgg ttgattcaat ccaagtctgt acccgtatca gcaactggct 1740
tgaattcact tcctcttttt ccctagctcc aacagttgag attgttgtcc aatctaacac 1800
tgaatctgaa tctgaatctg aaacttacca ctatattgag acaatcaatt tatctccagc 1860
tgtggaagca ttgcgactct attttcaaaa tggcggagga gcttgctata tctacccatt 1920
aaatgatgct gaagatgaat tggttctggc ggccatacca gaagtcattg aacagaaagg 1980
tgatattact ctgttggttt gcccggaact cgatctggat tacaaaacta agatctatgg 2040
cgcagtgagc tcactgttga atgataacaa agtgggctat ttcctgattg cggatagcaa 2100
tgatggagaa tctgtgtcag gagtatggaa tagtgctaag gccgccgcct attatcccca 2160
gttggaaact aacctaaaat tttccacgtt gcctggggat aaggacattc gtatcagcgg 2220
ttatcaggat gatgatgaaa cacataaacc gaaaaacttg gatgagctca ggacaatcaa 2280
cgaggcgttg gcacaggata ttgatgcaag attgctcgag gagaaacaac gtgctgtcat 2340
cattccgcca agtgctgcca ttgcgggcat ttattgccaa acggataatc gtcgcggtgt 2400
ttggaaagcg ccagccaacg ttgcgctcac agggatcggg agtttgcttg ataaggtaga 2460
cgatgaacgg cagggagaga tgaatgacaa gggaatcaat gtcatccgtt catttaccga 2520
ccgtggtttt atggtctggg gagcccgtac ttgtgtggac gctgccaaca tcagctggcg 2580
ttatattcct gttcgtcgcc tgttcaattc cgttgaacga gatatccgcc aggcgctgcg 2640
cgctgtgttg tttgaaacta atagtcagcc tacctgggta cgtgctaagg ctgccgttga 2700
tcaatatctt tatacccttt ggcagaaaaa tgcattgatg ggtgctcgcc cggaagaagc 2760
ttattttgtg caaattggtc aggatatcac catgtccgag gctgatatta aacagggtaa 2820
gatgatcatg actgttggtt tggcagcagt gcggccagct gagttcatca ttctgcaatt 2880
tacgcaggat gttgttcagt aatctccatg actaaacgcc aggcactgta ttgacagtgc 2940
ctactctaac catcttggag gaggtgatga tgatggagag actccaaccg ggtgtgactt 3000
taacagaaag tataatcacg atgggtcagc aagagatacc cagtgctgtg ccggtgttta 3060
ttggttacac cgttcgttat ccggaacaat cggaagcatc agtccgtatc gacagtttgg 3120
ccgagtatac cagcctgttt ggtgacgacc atgtgatgat gtttgctgtc aggcactatt 3180
ttgataatgg cgggcaacag gcatttgttt tacccctgaa ggacaatatg ccatcagtgg 3240
agatgaccac agctgaagcg gaaaatctga tagccgcatt gcgctctgct acggttagcg 3300
aagccattgg tgggcatagt cagattacac tgattttggt accggatatg gctcggctta 3360
atgacagtga tattgatgac tcctcaaccc aggtaagcct gtggtcccaa ggctgggagg 3420
cgctgctgca attgagtcag gttaggccca acctctttgt gctgttagat gcgccggata 3480
atgttgaaca ggcgcagaag tgtatgacaa cgctatcgtc agattatcgt caatgggggg 3540
cagcatattg gcctcgtctg gaaactacct atcagaaaga aatatctggc aaggacaatg 3600
aatctcaggg aattttccag gggactgttc tgtcacccac agccgcggtc gcagcggtaa 3660
ttcaacgcac ggataacgac gcgggtgttt ggaaagcacc ggccaatatt gccttatccc 3720
aggttattcg acctgttaaa tcttatcttc agggaagtgt actgtttaac agcagcggca 3780
cttcgctcaa tgtgatccgc agtttcccag gtaagggcat acgggtatgg ggatgccgca 3840
ctctggaaaa cacggataat acgcagtggc gctatctgca aacacgtcgg ctggtttcct 3900
atgtaacagc gcatttgacc caattggctc gcatgtatgt ctttgagcca aataatgaac 3960
ttacctggat gaagttaaaa ggacaaagtt acaactggtt acggcaatta tggttgcagg 4020
gtggcttgta tggttcacag gaggatgagg catttaacat tctgttaggc gtaaacgaga 4080
cgatgactga ggatgatgtt cgtgcaggaa aaatgatcat gaaagttgag ttggctgtgt 4140
tgtttcctgc cgaatttatt gagatcagtt tggtgtttaa tacccaaaca gaggcgctgt 4200
cttaagaagg aaaaagtacg atgaacgatt attacacacc cgtggtatcc catcgtttta 4260
tggcgagttt tatttttaac cgcattcccg atccgctgga tattcgtttt cagcgtatct 4320
ctggccttag tcgggaacta caggtgactc agtacagtga gggaggagaa aatgcccgta 4380
ataactattt agctgagaaa atccaacacg gtacgttgac tttggaacgg ggcgtgatga 4440
cagtctcgcc attgacctgg atgtttgatc gggtattgag tggtgaaaaa atcgcttatg 4500
ccgatgtggt ggtgatgcta ctgaatgaaa attcactgcc attgtccagt tggacgttga 4560
gcaatgcgct gccggtacgc tggcaaacca gcgactttga cgctaacagc aatgccatat 4620
tggtgaatac ccttgaattg cgttaccagg atatgcgctg gcttggagtc aaaatatgac 4680
agtagaaatc agagagttac ttatccaggc aaaggtagtg ccatcaacac gaccgactga 4740
atcagaacgg caaaaccatt ctttgataca ggaaagtctg gatgaggcga cttgggtgga 4800
aacgataaaa cgcgaagtgt tggccgcatt acgcgatgag gaagggtggc gtccatgagt 4860
ctgattgaac gtggtttagc taagctgaca attaatgctt ataaggatag ggaagggaag 4920
atacgggcag gaacgttgca ggccatgtat aaccctgact ccttgcaact ggattaccaa 4980
acggattatc agcaatccca agcgattaat agcgaaaagc aaagtagcat ttatgtacag 5040
gccaagcccg cagggttatc acttgaatta atttttgatg ccacgatgcc gggtaacaaa 5100
acccccattg aagagcagct catgcagctc aagcaactgt gcagtgtgga tgcaaccagt 5160
aacgagacgc gattcctgca agttaaatgg ggcaaaatgc gttgggaaag tcggggttac 5220
tttgctggca gggccaagag tttgtctgtg aattacactt tgtttgatcg tgatgcgact 5280
cccttgaggg tacgggtaat attggcatta gtggctgatg aaagtctggt gttgcaggag 5340
actgaacaaa atctgcaatc tccggcaaaa atcgcattac gcatacagga tggggtatct 5400
ctggctctga tggcagccag tacggcatca acattgtcag gcggtgtgga ttatctgacg 5460
ctggcctggc aaaacggtct ggataatctc aatgggttcg ttccgggtga aatattgcag 5520
gccaccaggg gagacgaatc atgagccacc aactgaaaat tattgcagat ggtaaggcac 5580
tgtcactttt ggccgcggta gatgtggaca cctgttatcg ggttaacagt ataccttctg 5640
cgacattgaa actgagcgta ccggataggc cactctcttc tttcagtcag acggatgttc 5700
agacagaact ggcccactgt caggtaggga aaaccctgcg tctggaattg attgatggta 5760
gcaaaaaatg ggtgctgttt aatggtctta ttacccgtaa ggctctgaga attaagaata 5820
agcaattatt gctcactctg gttgtcaagc atcggttgca actgatggtg gatacccagc 5880
attcacagct gtttaaagac aaaagcgaaa aagcgatctt aagcacgcta ttgaatcaga 5940
ccggaatcaa tgctcgcttc ggaaagatag cggcgttaga tcaaaagcat gaacagatgg 6000
tgcaatttcg ttgttcagac tggcattttc tgttgtgccg actgtcggca accggtgcat 6060
ggttgttacc tgccatagaa gacgttcagt ttgttcaacc tgatgctctg aaatcaaact 6120
cagcctatac cttgaagagc aggggggatg agaacaaaga catcgttgtc aaggatgctt 6180
actggcagtt tgacaatcaa atcaaccccg ctttgctgga agtcagtggc tgggatatca 6240
gtaagcagca ggtacaatca ggcggtcgct acggaaaaat cgcgttgggt aaggcggcac 6300
tctctcctga tggattggca tcccttaata aaacgggttg ggacatttgt tatagcagtc 6360
cgttaacaac ccaggaaagc ggttatctgg cacagggatt attgcttaac cagcgcattt 6420
ctggggtgac aggagaattt ttgctcaaag gagatgggcg ttaccagttg ggagacaaca 6480
ttcagctgac tggatttggt tcacagttag atggtacggc aagcattact gaggttcgcc 6540
accgtcttaa tcggcgaatt gattgggaaa ccacggtgag cattggttta caacatgaat 6600
atttgccgat attacctgat gctcccgaac tacatattgc gacagtagcg aaatatcagc 6660
aggacagtgc ggtgttaaac cgtatcccca ttattctgcc ggtactgaat cgtcccaatg 6720
aatttttgtg ggccagattg gggaaacctt atgctagcca tgaaagcggt ttctgttttt 6780
acccagagcc aggtgacgaa gttattattg gtttttttga aaatgatccg cgttatccag 6840
ttattttagg tgctatgcat aatccgaaaa ataaggcccc ttttgaacca acccaagata 6900
atagggaaaa agtattgatc gttaaaaaag gtgaagcgca acaacaatta gtcattgatg 6960
gcaaagagaa aatgatccga attaatgcgg gtgaaaatca aataatgctt cagcaagata 7020
aagacatttc tctgtcaacg aaaaaagaat taacactgaa agcgcagaca atgaatgcca 7080
cgatggataa atcattggca atgtccggga aaaacagtgt tgaaatcaaa ggcgcaaaaa 7140
ttaatcttac ccaatgaaag gtgacgatga atggaaaatc aaatactgac acaactctat 7200
ggtcgtggtt gggcttttcc tccggtcttt tcccttgaaa agggggtaga gatggctgaa 7260
ggggcggaag atgtgagaca aagtttgcag attctgttta gtactgagcc gggggaacgt 7320
cttatgcgtg aaaattatgg ctgcggatta aatgatttta tgtttgaaaa tatccgcaat 7380
gaacttattg ctgaaattga atcccatatc catgacaacg tattacgata tgaaccccgg 7440
gctgatatga ctgatattca ggttcgtcaa tcccctggca tggggaatac tttgcaagtg 7500
caggtcatgt atcgcctgag agggagtgat atcaatcaac aaatccaggg agtacttgca 7560
ctgagtgaag gccgggtgac ggaggtagta tgagtgaagc gattgtggtg gatggtgacg 7620
tgttacagtt tgatcccaac tttggcaatc ggcaggtgac ggttcccagc ccaggaaaaa 7680
ttagcggcac aggacatgcg caggtaagtg gaaaaaaagt gtgtattctg ggggatgaga 7740
aacaggtcag ggtttctgca acctatatta caacaacaca tactacgccg ggaacaggaa 7800
ccattactat cagtgctctg gatgctggcc agcaggccct tcagtgtacc agtggggcgg 7860
ctttaattat caaggggcag caatttacgg cgatgtttac gcctgaattg ccagccatga 7920
ataatacagt gactccgcca caaccggatg ttacgacacc ttcatcagga aaaggacgtt 7980
ttatcactca acaaaatttt gctaccgtaa attagagtat tgactgaatt aaatagaatt 8040
aacgaaggtg taaataatta tttatttgct gacgaatcgc tgtgacaaat aaacacaggt 8100
gatgttatgg aattaaatga gttaactaac aaattgtcaa atttggtgcc aatgaccgat 8160
tttaaattag ataatcgagc cagtttgcaa ttgcttaaat atattgaagc gtatacgaag 8220
ataataccct ttaattctgg cgataaatat tggaatgact ttttctttat gtcaggaaat 8280
acgccagaga aacttgcaaa attatatcag aaagaaatag aacccaatgg ggagttatta 8340
cctcagcagg cttttttgtt ggcggttttg cgtttattgg aaacaccaat atccttatta 8400
aatgtattac ctgctgctca tcgtgagctc tattatcggg agcttttagg cttgtcttcc 8460
catgcggcac agcctgatca ggttgcttta tctatggaac tgaattcgac agtgatggaa 8520
cagctgctcc ctgaaggaac cctgtttgag gctggtcagg atgaacaagg caatgcattg 8580
caatatgccc tggatgccag tttgctggct aatcgtggat atatcagtga cttgcgctgg 8640
ttacggaatg acggggaaaa gcaatgggtt acttctgctc catgggattt acaggcacag 8700
gtgtcactgc cgtctgatgg gatacgatta tttggtaaga caaatagtga tcagcaggta 8760
tttggtgggg tgttgataac gtcatcactt ctggcgatgg aagcggggat aaggaagatc 8820
attgttactt ttgagcagga gatgaacacc caagaactgg tggcacaggt cagcagtgga 8880
aatcaatggc taacattgac gtctgaggta aataagaaag aggtcacact gacactgtca 8940
gacaaagaac cggcaatcag tgcgccagag gatctggata atctcttttt cacgcaaccg 9000
gtactcaggc tacagggaaa ggatagtcag gcactgccgg aggtgacggg tatcagcgtt 9060
tcggaaaagg atgatactaa ggatacctct tttgagatgt atcacttaac accatttggt 9120
tatagcagtg atatagagcc attggaggaa aatccagcgt tatatttagg ctttactgat 9180
gtaaagccag ggcaaacact ggcgctgtat tggaaattaa aatccccgca gcaaccaacc 9240
gtttcctggt attacctgga tcaacataat caatgggctg aattggattc atgggtcagt 9300
gatggaaccc agaatctgta tcaggatggt acttggcacg ttgagttgcc tgtggatgca 9360
tccaatcagg cagagcagat gccagttgga cgctattggt tgcgggcagt ggtggaggta 9420
cccgctcatg agggggcgtt ggggaaggct ccttggctat atggtctaat ctataacgcc 9480
atgacggcaa ccttggttaa tgtagatagc atcagtgaca gccatttctt aacccctttg 9540
cctgccagca gcatacagcg gcccgttgaa cccatcattg tgttggcatc ggtcaaccag 9600
ccttgggcat catggggtgg acgtatacct gaatcctaca gtgccttttt tgaacggata 9660
gctcaaaacc tgtctcatcg aaaccggtcc ttaacctggg gaaatatggt gacattactc 9720
aaagagcgtt atgtcagcat ctttgatgtt aagtatccag gtaatgatga actcaccaga 9780
gtgccagcat tggagcagca gcaactaaca gtgattccag caaaccggta caacgatagc 9840
gatgattctc tgcgtccggt actgaatcct gctcgtctgc aagagatggc tgattggttg 9900
cagcagaaag actctccctg ggcctctatt gaggtcagga atccagaata cttggatgtg 9960
aaaatccatt acgaggtgat ttttaaacct gatgtgaacg aagattttgg ctatcgccag 10020
ctacagcagc aactgtgtga ggtgtatatg ccttggagca tagatgagca gcggcccgtt 10080
gtattgaata acagcattaa ttatttccag ttgttagcca ctattcaaca gcaaccgctg 10140
gttgagcgag tcactcgtct gacactacat cgggctgatt cttctgatga gagtgatggt 10200
acagcatctg tggaagccaa agataatgaa gtgcttattt tagtctggga agaggacgat 10260
aatctgcaat accgaggaaa tgactatgag taatcaggat gcactgtttc atagcgttaa 10320
agacgatatt cactttgata ccttgctgga acaagctcat caggtgattg aaaaacaggc 10380
tgaaaaactg tggagtgata cggcagagca tgatccgggt atcacatttt tgcagggaat 10440
cagttacggt gtgtcagatt tggcttaccg acatacatta cccctgaaag atttactgac 10500
tccggcgccg gatgagcagc agcaagaggg aatttttcct gccgaatttg gcccgcataa 10560
tacactgact tgtgggccgg tgacagcgga tgattatcgc aaggcattgt tagatctaca 10620
cagcagcgac agcctggatg gtactcagca ggatgagggg gattttctgt tccggagtgt 10680
gcaactggtg cgtgaaccgg aaaaacagcg ttatacctat tggtatgatg caaccaagag 10740
ggaatatagc tttgtcaaca gtgaaggggc taaagagttt accttgcggg ggaattactg 10800
gttgtatctg gaaccaaccc gttggactca gggtaatatt gccgctgcta ccagacaact 10860
gacagaattt ttgactaaaa atcgcaatat tggtgaatct gtcagcaaca ttatctggct 10920
acaaccggtt gatctgccac tgttgctgga tgttgaactg gatgatgatg taggtgcaca 10980
ggatgtcccc ggtatttttg cggcggtgta tagcaccgca gagcagtatc tgatgcctgg 11040
agcacagcgt taccgtacgg aagtactgca aaatgctggg atgagcaatg atcaaatctt 11100
cgaaggtcca ttattggaac atggctggat accagagctg ccggcagccc gtgattatac 11160
tcaaaggctc actctcaatc ttagccggtt ggtaaatagt ctgcttgaga ttgagggcat 11220
taaacatgtg aatcgtcttc gtctggatga tagcttcgat aaaactgcta ttgaacccgt 11280
taagggggat acctggtcgt ggtcgatcaa agagggctat tatccacgtc tttggggaga 11340
agacccactt aaccaattgg cgcaacaaaa tggcccgctt agggtgatag ccaaaggagg 11400
gattagcgtc agtgtgagta aagagcaaat ccaggccagt ttacccagtc aatcactgat 11460
tcaaaatgag ccggtaatat tggcttacgg ccagcaccgt gacgttggca gctattatcc 11520
cgtcagtgat actttgccgc cttgctatgg actacaacat tctttgtctg aaagtgaaca 11580
cttattgcca cttcatcaat ttatgttgcc atttgaacaa ttattggcct gtggttgtca 11640
acagatagcc atgctcccgc ggttactggc ttttcagcgc gaaggttatg aggtttgggg 11700
tgatcagtgg ccctttaagt caggctcagt gaatgatgac gcccatcaag attatgcccc 11760
tgcattaaag gatttgttag gacagattgc gctggatagt gatcatgaat tggatattat 11820
taattacttg ctgggttact ttggcacaca gcgggcaccg cgtaccttta cgacacaact 11880
cgatgatttt cgtgcggtcc aacagggtta tctggcccag caaccgacat tgacttacca 11940
ccgctccaat attcgtatcg atcaggtatc gtcgctacaa aaacgtattg ctgctcgcat 12000
ggggctgggc ggtgagttgt ttaaacctca accggatctg agccaactgc ctttttattt 12060
gattgaacat cgagcgttgc tgccagtcaa acccaatagt cagtttgata aggaacagaa 12120
accagcctcg gtgacagagg aggggggcag ccaaacaggt caacattatg tggtcattga 12180
acagaagggc attgatggca agctgacaca ggggcaagtg atcaatttaa ttctgtatga 12240
aggagagcag ggagaaaccc aatttacgat acgcggtcag atggtattca aaaccgaggg 12300
ggataagttt tggttggatg tgaataatag tgcgcaactg gaatataatc tggcgcgggt 12360
aatgacagca gccaaggcga gtaaactctt ttggcaaaac agcccggtat ggatggagga 12420
tatgggctat cgtctggcct atgctagtga ccaatcctca ttgcctgtga atcaacggcg 12480
cttgacccgc acagtgcaaa ctccattccc gccgatggtt gttgtaggta gcgaaatcac 12540
cctgttaaag caggtgggga tagtcaattt aaaaaaagcg gagtcagaaa aactttatgc 12600
aaaagttgtt agctttgatc gcattgaagg gaccttgatt attgagcgtt tgggtaattc 12660
cactctggct tttcctacct cggaagaggc gtggcggtat agttggtatt tttcggggga 12720
gaaatatgaa aggactgacc gcttttcatt tgtgattagc gtagtagtga acagtgactt 12780
aattaaattg cccggtgttg atccctataa attggaagaa tgggtgaaag aaacgattct 12840
taccgaattt ccagctcata tttctatgat tatccattgg atggatcggg aagccttttt 12900
aaatttcgcc aatacctatc agcgttggca aaataatggt acgccactgg gggatgcggc 12960
ttattccatt ctagaaagtt tgacacttgg taaattgcca tctgccttaa aaggtgttgg 13020
cacaatgcgt attgccacat ctagtcaaag agaagaagtg gtgggtagta atggtgatca 13080
atggaataca gatggaataa cccagaatga attattctat gttcctaaag agagctagga 13140
aaaataaata tctgccacta atgatgttga attaaatatg ttttctggag ttaatcatga 13200
acgaaactcg ttataatgca actgtacaag aacaacaaac attatctaat ccaaaagctg 13260
ttggacctga catcgataaa ttaaaggata aatttaaaga gggcagtatt cccctgcaaa 13320
ccgatttcaa tgagttaatt gatattgccg atattggacg taaagcctgt ggtcaagcgc 13380
cacaacaaaa tggcccagga gaaggattga aattggctga tgacggtacg cttaatttaa 13440
aaataggcac tttttccaat aaagactttt ctccattaat attaaaagat gatgttttat 13500
ctgtagatct tggtagtggt ctgactaatg aaaccaatgg aatctgtgtc ggtcagggcg 13560
atggtattac agttaacact agcaatgtag ctgtaaaaca aggtaacgga attagcgtta 13620
ctagtagtgg tggtgttgcc gttaaagtta gtgctaataa gggacttagc gttgatagta 13680
gtggtgttgc agttaaagtt aatactgata agggaattag cgttgatggt aatggtgttg 13740
cagttaaagt taatactagt aaaggaatta gcgttgataa tacaggtgtt gcagttatag 13800
ctaatgctag taagggaatt agcgttgatg gtagtggtgt tgcagttata gctaatacta 13860
gtaaaggaat tagcgttgat ggtagtggtg ttgcagttat agctaatact agtaaaggaa 13920
ttagcgttga taatacaggt gttgcagtta tagctaatgc tagtaaggga attagcgttg 13980
atggtagtgg tgttgcagtt atagctaata ctagtaaagg aattagcgtt gatggtagtg 14040
gtgttgcagt tatagctaat actagtaaag gaattagcgt tgatagtagt ggtgttgcag 14100
ttaaagttaa agctaatggc ggaattaaag tagatgctaa tggtgttgca attgatccta 14160
ataatgtact ccccaaggga gtgattgtaa tgttctctgg cagtactgca ccaactggtt 14220
gggcgttatg tgatggcaat aatggtacac caaatttaat cgatcgattt attttaggtg 14280
ggaaagggac tgatattaat ggagtgagta ctaatacagc ttcaggtact aaaaatagta 14340
agttattcga tttcagttct gatgaagcta cattaactat tgatggtaaa acactgggga 14400
gagcattatc gttacagcaa atacctaatc atgcacactt tagtggaata attatggata 14460
cagagaaagt taattattat ggaagtaaaa aaatcacaac aaatgtgtgg ggtgtaacaa 14520
caggagataa tacttcagta cgatatattt ataagtcatc aggtgtactt gactctaaca 14580
ataatgtctc caacagtacc ttaggcggaa acagtctgca gacgcacgat catgatatta 14640
agataacggg cacaggaaaa cattctcaca aaaacaaagt aacagtccct tattatattc 14700
tggctttcat cataaagctt taatatatat gaaaaattga aaatataaat tatccattaa 14760
taataaagag gatattagca tgacttcgga gccaaatctg ttaaaccgga ttacaattac 14820
tattgaagct aataatcaac aagtagctag aaaagtattg catggctcct tgcttaatca 14880
agctaatata aataaattat ttaattcata ctttaatgaa tatgaaatta ataggggtgt 14940
ttatttagaa acattaatcc tgaatcttgg tacgataaat ttccatgatt ttaattcatt 15000
gtttcctact ctcctaaaag ctgcattgaa taaagaattc agtcaatatc agataaacaa 15060
ccatagggaa gaaatgctat ttaatgagac aatatcaaat caagctactg ataagtctta 15120
catatttggc gataacaaat taattgatgc agagaatttc attcactttt tatatcaaaa 15180
gcattccaca ttaaatctag tagaagcaat gggaaataat ggtattgaaa aattaacaaa 15240
tcagttaaca caaatagaaa ataaatttgc gttattattg gcaaaaagtt gtttgtctga 15300
ggaaggctta aaacgactct tggctatcaa acaacccgat ttattaatcg ctatcaatcg 15360
cagattatct gaaagaataa atagaccaca atatcaggag aagcttgttt cctgcggaca 15420
actgatattt agtgctctgg gatatataca acagtacaat atacaggaaa ttcctaaacc 15480
ggatgaaaaa gttattgcac gcataacaac tgaacttaat aataatggtt tgcttaatac 15540
aatacctatt attacactat ttcgtcagag tgggattaac gattcatcac taaatgattg 15600
gctaaagaaa atctggcagg tgagatcaat ttcacagtta tgcagaaagt atctttctgc 15660
taaggaatac caatatctgt cagaacattt tgtttcaaag agcgtcgata aaaatagata 15720
tgatgaagag cccgtaaatc agagcatatt atcaaggttg aataataatt ccattaaaga 15780
aggaaataat cacagtcaac tctgtactct cagtagacta tattctgaac ccgttgtatt 15840
acctgaacaa accattctac gtcaggttag taatacagta gatcagagca tattatcaag 15900
gttgaataat gcctccatta aagaaggaaa taaccaaagt caacttcgca ctctcagtag 15960
actatattct gagcccgttg cattacctga acaaaccatt ccacgtcagg ttagtaatac 16020
aggtatatta attctatggc caatgctacc tacactattt aaccagcttg gtctacttga 16080
gaaaaagaaa tttatccatc gtcaggccca gtttaatgcc gttgattttc ttgattacct 16140
gatttgggga accgaagatg tgaaagtgga acgaaaggtt ttgaataatg ttctatgtgg 16200
gttaatggct gatgaaatta ctgaaccaat gcctattgaa ccagaaaaac aatggataat 16260
aattcaatgg ctggacgcta ttatctccca actttctggc tggaaaaagt taagtcgtaa 16320
tgacgtccgt caattatttc tacaacgacc aggagaatta ctgatcaatg aacaggaaat 16380
taaaatcaca atacagcaac aaccatttga tgctctgtta actgattggc cgtggccaat 16440
gaatatggct tgttttagct ggttgagtca accattaacc attacgtggt tataaccatt 16500
gaccacaatg acttagtctg agtaaaaaat atgaatatat cgcctgtttt ttatgattca 16560
ttgaatcagg ataacgaccg tgatctatcg tttttattta gcgaactgga acgaatagat 16620
ctcgctcttc aacaccattt ttattgtgta gaaagtcagc gaagtgagct cctggatgag 16680
tttctgctca ctgaggcgga agtggtgacc aggctggata agccacttgg taaacctcat 16740
tggataaatg atgattatct ggcgatatcg caaaagggca atgtaagcct aatggcagcg 16800
tccagattaa tggatctgat cgaacgcttt gaactgactg attttgagcg cgatgtttta 16860
ctattaggct tattgcccca ttttgatagc cgctattatc gactgttttc gctgattcaa 16920
gggggacaac agggtcgatt accttctttt gcgctggcat tggaactgtt ttgccactcg 16980
gcgctggaga aacaggtaca gcaagcgagt tttctgcacc gggcaccttt gatgggttgc 17040
cagctattat ccatcgatac tagtcaaaaa acgctggcct ggctccagac tccctttatt 17100
actgacagcg gggtatatca ctttttactg gggcatcact acattatgcc ggctttagaa 17160
cattgtgctg agtggttaac accgacaggg attggctgtt atcctgaagg attaaaacaa 17220
gtactgggta acgtattgtt atctgacaac gataatatta gaccgattgt cttattacgg 17280
ggaatggccg gcagtgccag agcttatacc attactaata tgatggcttc agaagggaag 17340
caaacactgc tggtagatat atccaaactt gctgatagcg atgaaaaaaa cattattctt 17400
cagataaagc atattttgcg ggaaacccgc atgcatggag catgtttatt attacggaat 17460
ttttgcttgt tagtggaaca gaataaacaa ctattggact ccctgtcaga gttattgaat 17520
caacctgaat taagaattgt ttgcctgatt gagccttatt ccccattggt atggctgaaa 17580
aagataccgg tattactgat tgagatgcca cttttaacgc ctgcggaaaa agccagattg 17640
ttaattgcca gcttaccgga taattgttcc gaggatattg atacgataac tttaagccag 17700
cgttacactt ttaacccaga aaccctgcca ttgattttgc aagaggccca gctttatcaa 17760
cagcagcgag atccgctgga tatcttgcag caatgcgata tacgccaggc attaaatttg 17820
cgtgctcaac aaaatttcgg tcaattggca cagcggatta ttcctaagcg ctcattaaag 17880
gatttattgg tatccgatga gattgctcag cagttacggg aaatactcat agcaattaag 17940
tatcgggaac aggttctggc gggagggttt aaagataaaa ttgcctatgg cactggtatc 18000
agcgccctgt tttatggtga ttcaggcact ggaaaaacca tggcagcaga agtgattgct 18060
gaccacattg gcgttgactt aataaaagtg gatttatcta cagtagtgaa taaatacatc 18120
ggtgaaacag aaaaaaactt atcccgtatt ttcgatttgg cggaacagga tgcaggggta 18180
ttattctttg atgaagctga cgcactgttt ggtaaacgca gtgaaactaa agattcccag 18240
gacagacatg ccaatattga agtttcttac ttattacagc gcctggagaa ttacccgggt 18300
ctggtcattt tatccaccaa taatcgtggt catttagaca gtgcttttaa tcgtcgtttt 18360
actttcatta cccgttttac ttacccggat gaaaaaatcc gtaaaaaaat gtggcaggaa 18420
atttggccta gaaatataaa aatatcggaa gatatcgatt ttaacgaatt agctcaacga 18480
acaagcgtga ctggcgcgaa tatccgcaat attgctttat tgtcttcatt ctttgcttca 18540
gagcagggga atgatgaagt cagtaatgaa aatattgaaa ttgcattgaa gcgtgaatta 18600
gctaaagtcg gacgattaac attttaaaag ttatcacaat gaaagtattg aaatattaaa 18660
taaatttatt accaaaaagt tatcacgata taatttaaga gaggtttttt atgttaaaca 18720
cgcaaactat tattgatgtc aataaggcaa tggatgccat gctgcgcgca tatctgaatc 18780
aagatattgc cattcgtttt gatctacctg aattggatac tatgcaatct gatgcgatgg 18840
taagtatctt tctttatgac attcatgaag atttacagct tcgctcggca gaatcaagag 18900
ggtttgatgt ttatgccggg aggttattgc ctggttgggt aaatattaaa tgtaactatc 18960
tgattaccta ttgggaagct tctaagccag cgactgatgc cagcagtccg gatagccaac 19020
ctgataacca ggcaatacaa gtgatgtcac aagtattaaa tgccttgatt aataatcgtc 19080
aattggcagg tattcctggt gcttatactc aggttgtacc gcctaaagag agtttaaata 19140
gcctggggaa tttctggcaa tcactgggta atcgcccacg gctttctctc aattattcag 19200
tgacagtacc tgttagccta aacgatggtc aggatagcgc gactccggtt accgcggttt 19260
cttctacagt ggaacaaacg gcatcgctca gtcaagaagt ggttagtcat gctttacgcg 19320
aattactcat tacggaatta ggaggaggag aggataaccg gttggtactg agtaaagttg 19380
aattatccgc agtgaaagag acgatgactc aagacagtcc ggctcagatg attatattgt 19440
tgtctgtttc aggcattaca cgacaggaat atttgaagga aattgataat atctttgatc 19500
gttgggtaaa taatgctgaa gttattacca ctattgatga ttgtgggatt agaattgaaa 19560
gtataacgaa agataatctt gtaggaattt aa 19592
<210> 94
<211> 18490
<212> DNA
<213> 光杆状菌(Photorhabdus)
<400> 94
atggccacaa ccacagttga ctatccaata ccggcttatc gatttgttgt ctccgttggt 60
gatgaacaaa tcccttttaa cagcgtttcg gggctggata ttacttatga tgtcatcgag 120
tataaagatg gcaccggtaa ttattataaa atgccgggtc aacgtcagtt aatcaatatt 180
acactgcgta aaggggtatt ccctggcgac actaaacttt ttgattggct taattccatt 240
cagcttaatc aggttgagaa aaaagatgtt tcaattagct tgaccaacga agttggaact 300
gaaattttaa tgacctggag cgtagccaat gcattcccaa cctcattaac atctccttct 360
tttgatgcca ccagcaatga tatcgctgtt caagaaataa aactgactgc cgatcgagtc 420
actattcagg cagcttaaag catcacgatg attgatatat cagacgggac aaaatgatcc 480
tcaaaatttg gcacaacggc tacccgtcca actaaattta ccctcttaca gttcacgcaa 540
aatatcgcac aatacaattg gaggcaatat gccaacaaca acttatcccg gcgtttatat 600
tgaagaagac gcctcactgt cactttccgt tcgctcaagt gcaacggcgg tgcccgtttt 660
taccgttgaa gatgacagtc aacttcatac tcctaccaga gtgaatagtt ggttagaata 720
tctgacaaaa aaagcagata aaaaattcaa ttctaccgac aaacttgata tcgcattgcg 780
cgcttatttt attaacggcg gcggatatgg ttatctcgtc aaagcgggtg aattaacaaa 840
tcaaattcca aaacttaacg atgtcacatt actggtcgcg gctggagaaa atatcaaaga 900
tgctgtgagt acactttgtc aaccgggcaa aggcttattt gccattctgg atggcccaac 960
cgaagagtta aagtctgatg gcaaatccag agatccgtat gatcaaagcc cttttgccgc 1020
cgtttattac ccctggctag ttgctgattg ggcagacaat attccgccaa gcgcggccat 1080
tgccggtatc tattgttcag ttgaccgtac ccgcggtgtc tggaaagccc cagcaaatgt 1140
catattacaa ggcggggtga aaccgaagtt taaagtcacc gatgacttac aaggtattta 1200
caacaccggt aaagccatca atatgatccg tgaatttccg aataccggtg tcaccatctg 1260
gggcgcccgc acacttaagg acgaagataa ctggcgttac atcccagttc gccgcctgtt 1320
taacagtgca gagcgagaca ttaaaaatgc catgagtttc gcggtctttg aacctaacag 1380
ccaacccacc tggaaagctg tacaccgagc tattgataat tatctctatg ccctttggca 1440
acaaggaggg ctagcaggaa acaaagctga acaagcttac tttgtgcaaa ttggtaaagg 1500
gataaccatg accgatgatg atatcaagca agggaaaatg attgttaaag tgggtatggc 1560
cgcagtgcgc ccggctgaat ttatcatcct tcaattttca caaaatgtag cacagtaacc 1620
gtactgaggc gcggtttaac accgcgtcca ttcagtctat tgaatggagg agacaataat 1680
gataacggag ataaaacagc cgggcgtcac catcacggaa aattcgatat ccccgaaatc 1740
agataatgaa tttatcggcg tccccgtttt tattggccat accgaaaaaa attcaagcca 1800
taaaacggct gttaaactaa atagcctgat ggactttacc caagctttcg gtgcatcagg 1860
attaacctat tattcagtac gccacttttt tgaaaatggt ggacagcaag cttatatctt 1920
gtcactgggg attaatcaac agctaaaaga ttttcaatca ttgattaccg ccctgcaatg 1980
gaactgggta aaacaagcca ttgccgcaga aaacgaaatc acattgattg ttgtgcctga 2040
tattacccgt tttaatgatc tcagcgctca aaaaagcctt tggctacaac tctggcaatc 2100
aatacttgaa ctgtgtaaaa gtcggcgtgg catcatggga ttactggacg cgcctgatga 2160
tccaacatta gcaactgagt gtttaaaaca attctcttcc actgatcgcc aatggggcgc 2220
cgtatactgg ccaaggctaa aaagtaccta ccaagaaaac ggtacataca ttgtactttc 2280
acctactgct gcggtcgccg ccgttatgca acgcaatgac agtcagaaag gcatatggac 2340
tgctcccgcc aatgtggctt tagccaacgt catcggtccg gtacgttctt acattgaagc 2400
tggaaccttg ctgaatcaag aaggcacttc gttgaatctg gtgcgtagct tccccggcaa 2460
aggcattaaa atctggggct gccgcactct ggataacata cctcattctc cctggcgtta 2520
tatccaaatt cgccgtttgg tttcctatat cgaagctcat ataacccaac ttggccgcgc 2580
ctttgtcttt gaacccaaca acgccatcac ctggatgaaa tttaaaggtc aggcccacaa 2640
ctggctacgt caattatggc taaaaggtgg attacggggc actcaggaag atcaagcatt 2700
tgaggtgtta ctgggtgtta atgaatccat gagtgaaacg gatatcttgg ccggaaaaat 2760
gatcatgaaa atcaggctgg cgctgttaat tccggcagaa tttattgagc tgagtctgac 2820
gtttgatatc cgtaacaata ccgtacctag ctaatctaaa caggggaaaa acatgtacaa 2880
cttatacacc ccgtcagtat ctcaccgttt tatcgccagt tttctgttta acaacattcc 2940
cagcccactt gatatcgcct ttcagcgtat atctggcctg agccgagaac tgcaaaccac 3000
ccaacatagc caaggtggag aaaacgccag aaacgtctgg ttatccgaga agatccaaca 3060
tggcagcctg gtgctggagc gcggtgttat gaccatcact cccctcacct tggtttttga 3120
tcgcgtgctg cgcggtgaaa aagccgtgta tgccgatgtt gtcatcatgc tactgaatga 3180
aaatgcgtta cccgtggcga gctggacagt cagtaacgcg ctaccggttc gttggtccac 3240
cagcgacttt gatgctaata gcaacaccgt actggtgagt tctctggaat tacgttatca 3300
ggatatgcgc tggttaggag taaaagcatg acggtagaaa ttaaagaact gattattcag 3360
gctaaagtca ccgattctac gagtgatcaa ctcgccccaa gaacattagc ccaagaaaag 3420
ctggataacg cccgtttgat tgacatagtg aaacgggaag tgttagaggc attacgtgaa 3480
ggaggccatc atgagtttaa ttgaacgtgg tttatccaga ctcaccctaa ccgcttttaa 3540
agaccgagaa ggtaaagttt ccgtgggtcg cttacaagcc atgtataacc ccgatacgat 3600
ccagcttgac taccaaaccc gctaccaaca ggatgaaagt gttaatcgtg ccagccaaag 3660
cagccgttat gtattatccc aacccgccgg attatcctta gttctgctgt ttgatgcctc 3720
gatgcccgat aataacatgc cgatagaaac ccagcttgcg accctgaaat ccctgtgtgc 3780
gattgatgcc agcaccaaag taccccactt ccttaaaatc aaatggggca aaatgcgctg 3840
ggaaaacaaa ggttatttcg cctgccgagc cagtagcctg gccgtcaact ataccctgtt 3900
tgaccgggat gccacaccat tgcgggccag cgccactcta tctctggtag cggacgaaag 3960
ctttattatt caagctaccg aacggcagtt aaaatcaccg ccggccactg cggttagcgt 4020
aactgatatg ctctccctgc ctttgattgc tttagatgct ggagcgtctc tggctggtgg 4080
cattgattat ctctcgctgg cctggcaaaa cggtctggat aatcttgatg actttacccc 4140
cggacaaaca ctgcaagcgc ggggggatgc atgaagatac ccatgataac cctcaaaata 4200
ggtggcaaaa cgctcaatca attgactgtc atcagtctga caataaacca tcaaatcaat 4260
ggcattccct cgaccaacat caccttgggg atcgctggcg atgcgagcca tattttcgac 4320
accaaagccc aagctgaact ggcaagttgt cgccccaata atgaactcac cctacagatc 4380
caaaaaaccg tggtgtttaa agggagcatc gttcgacaag cacttgaact gaaaggtcaa 4440
gacagcatca ttaccctgac agcaaaacat ccactacaaa agttaactca tagcctccat 4500
tcacaattat tcagtcaaca gagtgatgaa gcgattatca ggaaattatt caatcaggcg 4560
ggtatccaaa caacgataaa gcaggctcct caacttaaaa ccgttcatga acaaatggtg 4620
caatttcgtt gcaatgactg ggcattccta aaaagccgat tgattgccac taatacctgg 4680
ctgttgcccg gcaatgaatc ggttactttg ataacaccta aggccctgaa tcaatcgaca 4740
gtgcatactc ttcatcgaca ggccagtgct gaagatattg tgttatttgc agcggatctc 4800
caatggaata accaatatag ccctaaaacg gtgagtgtac gtgcctggga tattgctcaa 4860
caaaagcttt ccccagcaat taatacccaa aacagtcagc ttggcagtca taaattggcc 4920
gtggacagta tcgccgcact ggctgataaa gagtggcaat gggcttacag ctatccatta 4980
gataatgaac aagccaaaca ccttgctcaa ggcattatga ataacctgcg aagccataat 5040
atatctggca gttttgaaat cgaaggtaat caccgttatc aaccggggga tgtcttggcg 5100
ttaaatggtt ttggtcaggg gatggacggt caagggatta tcaccggagt cagtcagata 5160
attaatcagc ggcaaggctg gcacacccta ttaaccttag gcatgttacc cgatgtagaa 5220
ccgccggtgc ctcaggtgaa agagttgcat atcggtatcg tggaaaaata ccagcaagac 5280
cgccaatcac taagccgtat cccagtcaga atacccgcat taaacttgac caaaggtgtc 5340
ctttttgccc ggctaggtaa accttatgcc agtcatgaaa gcggattttg cttttatccc 5400
gaaccgggag atgaagtgat tatcggattc tttgaatgtg atcctcgttt tccagtgata 5460
ttaggttcca tgcataatcc gaaaaataaa ccaccgttag aacccagtga aaaaaatccg 5520
gtgaaaactt tagttatcaa gcaaggggat aaacaacaag cattaatatt cgataataaa 5580
gaaaacacgg tggcacttaa tagcggcgaa aataaagtct ctctgcaaca ggataaaaac 5640
attacgctca attcaactaa aaatctcatc actcaggccc aagaaattaa tatacaagcg 5700
gaaaaatctc tgtcagccac aggaaaatct ggcgtcgata ttaagggcgc gaaaattaac 5760
ttaacccagt aatgaggtat tgaaatgaca agccaaatat tagccaatat ttacggttgc 5820
ggctggaaat ttccgccaca gttttctatt gaaactggcg tagaaatggc cgaaggtgcc 5880
gaaaacgttc gccaaagtat gaaaatcctt tttttaactg aacccggtga acgaattatg 5940
cgtgaagatt atggttgtgg tctgaatgat tacatgtttg aaaatatcag tgatgaatta 6000
ttatcggaga ttcaaacccg cattgaagaa cgagtattgc gctatgaacc ccgtgctgaa 6060
atcacagata tccaagtaac tcagaaaaca gactcaccga atactttaca tattcaagtg 6120
acctatgccc tgagaggcag ccaaatcagt caacagcttg aaggggttct tgagatcaac 6180
gaaggtcagg caaaggtgag tctatgagca aacaactcat tattgatggc gacagcctgc 6240
tattcgagcc attattcggc aaccggcagg tcactatttt gatgccagcg accatcagag 6300
gcagcggaca cgcgcaaatc caaggcagaa agatagcgat tgtcggcgat gaaaaaaagg 6360
tacaacttca agcgcaatac attaccccaa gccacccggt acctggcata ggcacagtta 6420
ccattgctca attagatacc agccagcaag tcaacttttg ccacagccct gccacagtga 6480
tagttgtcgg gcagcaattt accgctcgat ttaccccatc acagccggca attaatccgt 6540
caaccgggcc agatgtcaca acacccagta tgggcaaagg ccgttttatt gccagtcaac 6600
atactatcaa cgccggataa ataactctgc aaaatcatta ttcaataacg ttcctattct 6660
gcaatagcta tcagcaatat attcaaataa caggtggtat aatatgggac tcaccgaatt 6720
aaaaaataaa ctctctgcta tcgtactcga tacggatttt aaacttgatg aaagaagtac 6780
actggatatt ttaaactggc tacaagaata tgctaaaaaa atccctttca atcaagagaa 6840
aaaacagttc tgggatagtt tctattttat tcaggaaaat agtcctgaga aattagccga 6900
tctttaccaa aacgttaata aaacgaatgg ccatttaccg gcccatcaag cttttgtttt 6960
agccttttta aaacttttag aaaccaccaa agtattattt aatacttttc cggcacgaca 7020
tcgtgatctt tattaccggg aattattagg tctaaaaccc agaaatgccc aagcagatag 7080
tgttgcttta ggcattacct taaatacaga taacacagaa catcttattc ctaaaggaac 7140
cttgttcgat gccgggcagg acagggccgg aaatccgcta caatacgcat caaatgcaga 7200
tttactggcg aatcaaggaa aattgagcga tctgcgttgg tgtcgaaaag ataatgatag 7260
ctggcaatct gcaatactac tgaaccactc agataatatt gaattacctg aaaacagtat 7320
tcgacttttt agtccaacgc cggatgatat tcccgtttta tccggttatt tgataacttc 7380
gtctttattt gctatgccaa cgggggaacg cagtattaca ttgactttag cagataattg 7440
gcatggtgat attaagcaca tcaccgctaa aatcagttcg ggagatcact ggctttcact 7500
atcagtaaaa aaagaacaag acaatagtat tcactatctt aaactttatt tatcaaccaa 7560
tgatgacccc atcggtcctc ctgatgcttt ggataatata gcgtttgatg taccggtatt 7620
aaagctgggc actgttcagg gacctatact acccaagatt acgggtattg aaattagcat 7680
taacggcaac agtaatgtac attattcctc tgataacggt attgaaaaaa tagatgcagc 7740
tagttttccc tttggacaat caccgtcacc aggttccggt tttaatctga ttgcccctga 7800
atggtatggt acagaaagcg ccaaaattac tcttactcct caatggactg gattacccaa 7860
agaggggttt aaagagtggt atcaaggata tagttctacc cccgaaaata atgcatttaa 7920
agtacaggct tatttaatca cacctcaaaa gagagaaaaa tttaatgaag ctcagtcatt 7980
atttaatgaa agtaaagaca agaaaccaca aggaaaaagc ctaactttta ccttacctgc 8040
aatggattat tcctttgcaa acagcccatc atctaataac tggcccgcat caatacgcat 8100
agaactaacc gaacaggatt ttatgcatgc ccaatattgg caaaatccta cgggtaaaaa 8160
acagccctat acccccaaaa tgaacacatt acaaattcag ttcagtgcca aagttaaacc 8220
cgaacaattt tccgtttatt ctctcacgcc ttttggttgg ggaaaaacag gagaaaatag 8280
aacatcatta acccatgata cattctattt aggttttacc gatgtattac caggacaaac 8340
tttatccctg tactggcagt tagaaggtat taaaaagctc cctttatcct ggtcttatct 8400
gaatcaagaa aatacctgga gtccattgga taatcaggtg catgaccaaa cccacaacct 8460
atttgatcga ggaatctggc gtacctcatt gccacatgat gcttcaaacc aagcctctca 8520
aatgccaaaa ggacaatatt gggtgaaggc acacatttta caaacgaatc aagcaaccct 8580
gactgatctg tattggtatc gaaaagataa tgatgtctgg aaatccgcaa cacctcttag 8640
cctttcaaat aacatgaaat tacccgcaaa cggtattcag atttttagcc caacatctca 8700
tgatgttcca gttcgatacg gctacctaat tacttcatct ttattctcat tcctcaagaa 8760
aggacgcaat atcacattaa ttttagcagg agatagctgg gagggtaatc ctgaaaacat 8820
caccgctaaa atcagttcag gaaatcactg gttaacacta tccgtcgaat atctgagtaa 8880
tactaatagt cttaagttgc aattatcaga taataataat gatcccatca gcccccctaa 8940
tgctctggat aatatgacgt ttgacacgcc attgttaaaa ctagaagcca ctcaggattt 9000
cactttgccc tggatttata aggtatgcgt taatagcaac aatatactct ctacctctga 9060
cagctcagat gcagcgatta ctcgtttccc ctttggccaa tcaccatcgt tgggttccag 9120
ctttagtccg aaaatcgttt tcccggaatg gtttgaatct gaatacgcat cagacaccac 9180
gatcacgatt acccctcaat gggttaacct gcccacagaa aacttttcat cgtggtatga 9240
cggatatatt aataaacctg ccgataatag cgtatttaaa atagagggtt atttacttac 9300
tcattatcag ggaaaaatca aactcacaga agctgagaca ggaagcgaaa cccaagcatt 9360
attcaatgga aacaatgcac cacaaggaaa aagcctgact ttcactttac ctaataggta 9420
taacttctat ccgcgcaacc atcagtcaat gaagatagaa ataaaactcg ttaaacaaga 9480
ctttatgcac actcaacata agagcaatcc cacaggcaaa aaaccaccct ataccccgca 9540
aatcagtgcc ttacaggtgg aattcaatgc tacagctttc catcgaaaat tctccgttta 9600
tcctctcacg ccttttggct ggggcaaaac aggagaaaat agcacaccat taattcatga 9660
tacattttat ttaggcttga ccgatatatc accagagcaa actttttctc tgtattggca 9720
gctaaagggc cttaaagagc tacctttgtc ttggttttat ctaagtgaag aaaatagctg 9780
gaaatcatta aatagatcaa cttacaacca aacccacaac ctgtttgaat cagcagaaca 9840
aagtatccta ttaccacggg atgcttcaaa ccaagcctct caaatgccat taggacggta 9900
ttggctgaaa gcacagatag aacaggagaa aaaacagata aagatagcgc ttcctgatta 9960
ttatccaaga atcagggggc tgttgtataa cgctaccatc gccactttaa tcaacgctga 10020
agctgttgag caatctcacc ttatcaacgg attggctgct aacaacatta aacaaccggt 10080
taactcatcc gttgccatca acgaagttat tcaaccctgg acatcctgga acggtcgccc 10140
aaaagaaacc gagtcagcat tcctggcacg agttcctgcc cggctctctc atcgtaaccg 10200
agtgctaagc tggggtaaca ttgccacttt attaaaagag aattttagta gcttattcga 10260
tgtcaaatac ccttctgtca gtgaattaac caaaattcca gcgccagaaa agcgacaatt 10320
aaccatcatc cccgacaacc gctataaaga taatgatgat tcactacgcc cagtattgaa 10380
ccaagccaga ctgaccgaga tggtcgaatg gttagatcga ttaagtagcc cttggacaac 10440
tattgaaatt aaaaatccca catatgttaa cgttctgatc cactatgaac tgatatttac 10500
ctcggatgtt aaccccgatt atggcctcca tcagctacaa caagaactca gtcgaaaata 10560
tatgccgtgg ggagaaaatg cagctattgg cgtaacaccc ggtaatcgta ttgactactt 10620
ccagttatta gcctcaattc aacaatcacc gctggttgaa cgggtcacca acttaacgtt 10680
aaaaaaaggc agccagccta ccgtaagtga aagtatagaa gccgccgatg atgaagtact 10740
gattttagtc tggtcataaa aacttcccca acctaaggaa ttaacaaatg aataatcgag 10800
atatgctatt tcctatcatt aaagacgata ttacctttga ttctttattc gcccaggcaa 10860
aagccgttat tgaacaacaa tcggggcagc tctggaataa tacaggtgaa aatgatcccg 10920
gcattacttt attagaagcc tgttgttatg gcgcatccga tctggcctat cgccacacat 10980
tgccactgcg agatttgctt actcctcaag aaaatgaacg aatagatgat ggcatttttc 11040
ccaaagaatt tggtccacaa caaatactga cctgcggccc aattaccgcg gaagattacc 11100
gtcgagcttt gttagatttg cgtagtgata acaccgttga aggttatttt ttctttaatg 11160
atgcacagct cattcgtgaa ccggaaaatc aacgctattc atattggtat aacaaagaaa 11220
aacgcgaata cagttttact caagaccaat acagcgaaca attacagtta acactgagag 11280
gaaactattg gctctattta cttcccagtc ggaaaaccca gctcgataac accctggctg 11340
aagaaagact caacattttt ctgaaagata accgaaactt aggagaatcg gtcagtaaaa 11400
ttatttggct agaacccatt aaactgtcat tgaaaattga tattcagctt gatgatgacg 11460
ccaaagatat tgctgatata tttgctaaag tttatatgat tgcagaacaa atggtgcttg 11520
aaaaaccatt acgttatacc actcaagcga tgaaagaact gggttacagt caggaacaaa 11580
tatttgaagg cccttattta caccacggtt ggataccgaa attacctcaa accaaagatt 11640
atactcaccc taccgtatta aatctcagtc ctttaattaa tcagttactg gctatcaaag 11700
gggtgaaaca tattacccaa tttacattgg ataagcctga taaaaaaatt tctaagttac 11760
caaatgataa ttggtcttgg gaaatcgctc cgggatatta cccaaaacta tggggagata 11820
ctccattaga attaattacc tcaccaacaa gcccactcac catcacggca aaagggggaa 11880
ttaaaattgc tattactaaa caacagatag aaaaaaacat aatgacagaa ccactaatta 11940
atacacagcc agaattattg aactggggta aacatcgcaa agtcctggat tactatccga 12000
taagcaataa attacccgct tgctatggat tacaaactaa tacccaacaa cagctacagt 12060
tgcatcaatt tatgctgcct tttgaacaaa tgctagcgaa taactgcgct gaacttgctt 12120
tattgccaag actattagct tttaaacaac gaggaaatac ggtacatggc attcaatggc 12180
cttttaaaga aaatacggtt ggtcaacatg ttcataagga catagtatct aatttaaaca 12240
ataatgctac gaaaatcgat aataatgccg atgactacga caaggaactc gttattctag 12300
attatttgtt aagatatttt ggggctcaat gtgcaatccc acgactatca ccagacccac 12360
cacaatcatc attaacagaa cctcagacta aaaaagattt tctatctact cagcgcgaat 12420
atctggctca acagccaaaa ctgacttatc agcgtaacaa tattcggatt gataaagtat 12480
cagcactgca aaaacgtatc gctgcccgat taggtctggg aggagaatgt ttcaaagcag 12540
agcctgactt agctcacctt cctttctacc tcattgaaca tcgtaggctc ttaccagtaa 12600
aacctgatat aaaattctat attgagcaac aacctaattc tctggaaatt gaaaatgata 12660
aattaaaaat cacacagaaa gattcagcgg gtcggttact gcaaggtcaa gttattaacc 12720
tggaatttcg tgagggctat gatgaattta cattgctaaa cttaatgata actgaagtga 12780
caagagatac attcaccatt agcattaata atagccgtga tctcagagac aatctggaca 12840
aagtgcaaca cgcgtttgaa caaacgaata atctgagctg gcacaatagc ttaatatgga 12900
tggaagatat ggattatcaa ttggtttatg ccaatggaga acaactggaa aaagcggaaa 12960
atgaacgatg gattaccatt aacaatcaaa gtgctttccc tgctatgatc ggagagaatg 13020
atgaaatcac actaaaaatt caatccgatt atgaacttaa aaccaaagtc gtgcggcttg 13080
attataacaa caaaaaaatt ctgattataa aagatgcgac atcaataaat aattttccgc 13140
caaaaagaga agcatcatat tattcttgct cttctctaaa agacaatggg tacggatatt 13200
cggatgaata taaatatgaa cttacttata ttgatacaga ttctacaaaa gaaaatgagt 13260
gctggattac tatcagcgat ccaaataatt tgttttctcc tgatatcatc gcagagaatg 13320
acgaaattat attgaaagct aaccctaatt atgagtttaa aacgcacgta gtaaaatttg 13380
atcgtattaa tagacaaata ttacttagga aaaatacaga cctggaaaat aattttccat 13440
cagaaaacaa cacatcgcac tatcgctggc atttctctgg tgaaaaatat gcccaaactg 13500
accatttttc atttgttgtc agtgcagtac tgaatcgaga attaattgag aggggcacag 13560
tcgatctcta taaattagag tcttgggtaa aaactgagat tttatctgaa ttacccgcgc 13620
atatctcact cgttattcat tggctatcat cggaagaatt cgaaaaattt gccagtactt 13680
ataaagtttg gcaaaataat ggcgctcctt taggtgatca cgcatataaa attctagaaa 13740
cattaacact tgggaaaaaa ccttctactt cagcaagaag gtccagcagc tatatagaag 13800
cacagtaata attcttacag aacattaacc catatttatc ttataatatc aaacatcata 13860
aaaacaatct tcagctcatt ataatgacat atttcatact caggtttctt catatctgtt 13920
aattacaaag agaatattaa tatgatctca gcaccaaatc tgttaaatcg gattatcatt 13980
actattgaag cgaataacgc acaggcagct aaaaaagtat tgcatggctc cctgcttaat 14040
caatccagta taaacaaact ctttgattca tactttaacc aatatgttgt taatcagact 14100
atctacctga agacactcac cctgaatctt ggcgaaatac gattaaatag ttttaattca 14160
cagtttgtta ttcggcttaa tactattctg agtcaagcat tgagccaata tcaggtaaat 14220
aatcaaactg atattgagaa atttatttat tacttatatc gaaaagattc tatattaaac 14280
ccaatagagg aaatcaataa tcgtgaaatt actgacatca atattaagca attaattaac 14340
caattacccc agatacaaaa caattggaca ctattattgg caaaaagctg tttatccaca 14400
catagcctga aaaaactcct ggctatcaaa aaaacagctt tattaaccgc cattaatcgt 14460
aaattatctg aaaagatcaa tatatcaccc tatcagcagg aatcggtttc cacctggcaa 14520
ttgatactga atgcgctgaa atatatacag cgacataata cacaggaaat acctgaaccc 14580
gatgcgaaag tcatatcact cattacaacg gaactcaatg acaatgccat taatacagca 14640
ccaattattg cattatttcg ccaagttata accaaccatt ccccactgaa taagtggctg 14700
gaacaactgt ggcaaacaaa gcgaatttca cagttatgta aaaaacagct gtcaattgaa 14760
gaataccaac atctatcgga gcgctttatt gccaaacacg ggaataaaaa taaatctgat 14820
aaaaaatcat ccatgacttc cgaaccgctg ttattacctg aacaccctcc accacgtcag 14880
gtcaataatg ctggaatatt agttctgtgg ccgatgttac ctactctatt taaccaattc 14940
ggcctgtttg aaaaacaaaa atttattcat cgtcaagctc aatttagggc tgttaatcta 15000
cttgattatc tcatttgggg aaacgaagaa acacagacag aacgaaaaat attgaattgc 15060
gttctgtgtg ggttaattgc cgatgaggac acggaatcaa tccctattga gccagaaaaa 15120
caacaggtaa tagaacaatg gttagatgca gttatcagtc aacttcctgc ctggaaaaaa 15180
ttaagccgca atgatagccg ccaattgttt ttacaacgcc cgggggaatt gctgacaaat 15240
gagcaggaaa tcaaaattac ggtacaacct caaccattcg atgcactgtt aaataactgg 15300
ccctggccgt taaatatcgc caaacttccc tggctggatc gccctttatt aatcaactgg 15360
taaaacattg acaaggttta tatgaaagaa catcaatata gaatagtcga tctacgctgg 15420
atttattccc atttggagcg catcgatctg ctgttacaac gtcactatta ccaaaagaga 15480
gacaaatacg attcattgcc agaaagtttt ttgcttgaag aagatgaatt agaacaacgt 15540
ctagcaaaac cgttgggtat tcctcattgg ctaacagcaa ataccggcgc tggtgataca 15600
gaaacagaaa atcattctgc ttccggcaca ttatcactgc tagtcacgcg ttttaaactc 15660
actgaatttg aacgtgatgt gttattgcta ggtttattac cgcattttga caaccgctat 15720
catgcgttat ttgctactct gcacggtaac agtaaaaaac agtggcccag ttttgattta 15780
gcgattgaat tatttagcca acatcaaagt aactggcaat tatttcaaca ccacttttta 15840
ccgcaagctc cattaatcaa tcaccattta ttacgactca ataaccaaga ggaacccatt 15900
tggctacaaa ctcaattttt aactcacaat gcagtctggt cttttttatc cggtcagcgc 15960
gtcattttac ctcccttaat atcctgcgct tactggcata ttccaacctc acagacttgg 16020
tatccaccaa tccttggtca tgcatttgaa aaaatattgc tgaatgaaac ggacgaaata 16080
cgcccgctgg tggttcttaa aggaaaacag gacagcgcca gagaactggc agtcagtaat 16140
attatgggaa ttcacggcat taacacttta acgttcgatt tatttcacct gccagatgaa 16200
gagtgcacca cctcaatact caatctgcta atagatgcaa tacgagaaac ccggctacat 16260
aatgcctgtt tattaatccg taacttttct ttgctggcag aggaaaagag aatatcgcat 16320
agagaattat cagctctact gaatcaaccc aaattacgtg tggtttgtct ggcagagtca 16380
gaagaatcat tagcatgggt taaacacctg ccgatagtgc aaattaatat gccaccggcg 16440
acgctggcag ataaaaaaac gatgctggaa gccagtttgc cagataatgt cactaaagga 16500
attaatataa ctcaattatg tcaacgtttt tcatttacag cagaaacatt accgttaatt 16560
atcaaggaag ctcatcaata ccaaatcctc cgacaaccgg aagatcaatt gaaagaatct 16620
gatctacgta aggcattaaa ttgccgcgcc caacaaaatt tcggtaaatt agcccagcgt 16680
atgacaccaa aacgaagttt taatgatttg gttatttccg ctgacttaac tcaacagttg 16740
aaagaaatca tcgcagcaat taattaccgt gaccaaattc tgggcgcagg ttttcgggaa 16800
aaaatcagct atggtactgg tattagcgcc ctattttacg gtgaatccgg gacggggaaa 16860
accatggccg cagaagtgat tgccagctat cttggtgttg atctgattaa ggtagatctt 16920
tctaccgtgg tgaataaata catcggtgaa accgaaaaaa atatctcccg tattttcgat 16980
ctggccgaag cggattccgg ggtgctgttt ttcgatgaag ccgatgcctt attcggtaaa 17040
cgcagtgaaa ccaaagatgc ccaagataga catgccaata ttgaagtttc ttatttatta 17100
cagcgactag aaaattatcc gggattagtg attttagcga ctaacaatcg caaccatttg 17160
gatagtgcgt ttaatcgccg ctttaccttt attacccgct ttacttatcc cgatgaagca 17220
ttacgcaaag caatgtggca ggcaatttgg cctgaacaac ttaagttatc agatcaactt 17280
gattttgagc atttggctaa acaggcaaat ctgaccggtg ctaatatcag aaatattgcc 17340
ttattatcat caatattagc tacagataat aatagtgatc aaattgaaaa taaacatata 17400
gcgcgagcat tgatacttga attaaataaa acgggccgat tgatttttta atcatttata 17460
cccaataaat ttcgagttgc agcgcggcgg caagtgaacg aatccccagg agcatagata 17520
actatgtgac tggggtgagt gaaagcagcc aacaaagcag caacttgaag gatgaagggt 17580
atatagaatt ggagtgaata tgacaaatat aattaaccct aataatgcga ttcttgaagt 17640
taataacgca ttaaatgata ttttatctca gtatttaact aatattgata tccgctttga 17700
tctaccagaa ataaattcaa tcccatcaac ccctacagtg agtatatttc tttatgatat 17760
acatgaagac ctacaattac gttctgctga accaagaagt tatcatccta ccaccagctc 17820
attattgccg ggatgggtaa atattaatta taactattta attacttact ggcattcaag 17880
taatccatca agcgacagtt ctacccctga tagtcaaccc aataatcaag cggcacaagt 17940
catgactgct attttaaatg cattggttaa caaccgacaa ttacctaaaa ttcctggcgc 18000
atataccaga gtcattccac ctcaagaaaa tctaaatagc ttaggtaact tttggcaagc 18060
gcttggcaat cgccctcgcc tttctttatt atattcaatt accgcaccgg taaaactgca 18120
aaatattaaa gatgtcataa agcccattag ccaaatttcc acttctgtgg atcaaaaatc 18180
aaatctggat aattcgcaaa tcaaccaagc cttatttagc aaattgggtg ccgatttagg 18240
tggcacacaa gatgttcgtc ttgctcttgc gaaagtgaat ctgacaacca aacctgctaa 18300
agaaaataat gaaaatcaaa ataataaaaa tgtaattatt gaagtttctg gcattaccca 18360
tttggattat ttacccagaa taaaaggtat tctttcaaca tgggtaaata gtcatagtgc 18420
tgttgttagg ataaatgata ttggtattat tgtttcagaa tataaatatg ataaattaac 18480
aggcgtttaa 18490
<210> 95
<211> 22492
<212> DNA
<213> 光杆状菌(Photorhabdus)
<400> 95
atgaatacag ctcaagaaat tattaaccgt ttatcgggga gagccgttac gcttggttgg 60
gatgttgtta ttgcttatga ccgaaaaaaa attaacactc tgttagagca acaatatgtt 120
gaaaaggtaa aaaacgggga gaacttcccg cttatcaact gggagaacca gagaaaaaca 180
cttcaattta aagatcttca attaggtgtt ccacttattt cttttgagaa ttcaacactg 240
gaaaattcaa gggcgcttgc cacgatagaa tttatttcag gagctattat tgaatttagt 300
gactccgggc aaataatcaa ctataagaag attgaaccta gtcatggtta tggcatggtg 360
ctgactatcg atctcatggc tggtacaggt tcagtagaag aacaaggtcg ggtgataata 420
aatcttaacg aaggcgccat actcgatttg catgttatcc aacaaccgcc agcagaagtg 480
gtagaatttt tccgcacttg gttgatggct aataaaatga cttatgaatt aggtaagctg 540
gatctgagta gtcaagctgg tctagtgcct cgttcttttc gtattcgtac tcagcgggcg 600
cctgaaaaaa ttcgtaaagc gacgagcgat gaaggaaatg gcgctgtttt gttgtttgtt 660
gccactaact ataaccctac aagtggaact ttacctgcca aggattatcc gtggctaatc 720
cctgaggaat attcaggcgc attgcttatc ggtaataaat gcttatttaa agacattctg 780
aaaccgaatc tggatcagtt gtttgataaa ggggaatgga cattaaaagt tcagcaaacg 840
gattctgatc aactgctgca ttatctggag gcaaactctg catatataac agataagcct 900
tatatggcag actttgaagg aactcaggat ggagtctgga caggacgtta taaatttgag 960
actggccggg gacattatgg ggtgtatgaa aatgtacgct ttcctatcaa tggaatgttg 1020
atgaaaccgg ctaaaactgg attacagtta tcaatagatt caccacaaag ccatcaattt 1080
aatgttgatt tcggaatgaa gtggttccat tgtgctaata taatgtgtgg ttattcctgg 1140
tttaacgaga cttacccatt ttatcttgat ggaaaatcat tttatcaagt tcatattgac 1200
cctgataaag aggtgattta ttttactggg ccagatgaag atattaatat tgtaggaaat 1260
tacagcccgc ctgcgtggtg gcaatctaaa tggcaaaaac atatcagtga tgattttacg 1320
gatatttcct cggaaaaatt taagcgactc agtcaaataa aattgccaga aatatgcatg 1380
tttgccgtga accatttatt atttcctggt cataatactt tgctgttgaa agacgtttat 1440
ttaccgggtg atatggtgat tttcggtgat attaacccat cacttaccgc ttttcgggtt 1500
acgccattaa aagcaacagt ggtggcaaag ggaacccaac aatttaaagc catagaaact 1560
aattgatgat tatacccttc atccttcaag ttgctgcttt gttggctacg ttcactcacc 1620
ccagtcacat agttagctat gctcccgggg attcgctccc tggccgtcgc gatgcatctt 1680
gaaatccata gggtatatat ttaattggat aagtcttttt tattttaaca ttataacctg 1740
attctttttg gataaaatta aaggattatt aacatgtcta ttacacaaga acaaatcgct 1800
gctgaatatc ctattcctag ttaccgtttt atggtttcta taggagatgt gcaagtccct 1860
tttaatagtg tttcgggatt agataggaaa tatgaggtta ttgaatataa agatggcatt 1920
ggtaattatt ataaaatgcc aggacaaata cagagggttg atattacact tcggaaaggc 1980
atattctctg ggaaaaatga tttatttaat tggattaatt ccattgaact caatcgggta 2040
gaaaaaaagg atattacaat tagtttaact aatgatactg gcagtaaagt cttaatgagt 2100
tgggttgttt cgaacgcctt tccgagctca ctgacggccc cttcatttga tgcttcaagt 2160
aatgaaattg cagtacaaga aatttcatta gttgctgatc gggtaacaat tcaggttccc 2220
tgataactaa aaactttaag gaaaaataat gtctgtacaa acaacttatc ccggaattta 2280
tattgaagaa gatgcatcat tgtctctatc tatcaataat agtccaacag caatccctgt 2340
ttttatcggt aaattttaca acttggatgg ttccttacct aaagtgggaa catgttctag 2400
aattaccagt tggttagatt tcactaaaaa attttcggta gctcctcctc aaaccatttc 2460
attgatcgcg tcgccaattg ctgacacaca agaaagtgta cccaaagcag ttcaatatac 2520
ttataaggcc gagtttgaaa cctcagaaaa tctggcaaat ggtgcctatg cggtacaaca 2580
ttatttccag aatggcggtg gtatttgcta tatcatacct ttagttagcg tgaaaaaaga 2640
ggatgctgcg attgagttaa caaaattacc tgaattaatt gaaagacaac aagagattac 2700
gttaatcgtc tgcccggagg acgataagac gctcactgtt gatagcagta aaaaatcgga 2760
tgtttataac agcatcaata cattattgag taataaggta ggttattttc tcattgcaga 2820
ttcagatgat ggcaaagcag ttcctgatac gttgccggaa aaaactgcgg tctattatcc 2880
tggtttacta acttctttta cacaacgcta tgcccgacct gccgattctg ctatcaaagt 2940
gaccggtatt acaaatatat caactctggc tgatattcac accaacttgg ccgatgacta 3000
ctcaacagca agtcaggtta ttaatgatgt tttggaaaaa aataataagc tcgcatcgtc 3060
tcccattatt ttacctccca gcgccgctgt tgctggtgct tatgccgctg ttgatgtgag 3120
tcgtggtgtt tggaaagcac ctgcgaatgt gatgttaagt aatgccacgc caatcattag 3180
tatttccgat gcggaacaag gtgtgatgaa cccattaggt attaatgcta ttcgtagttt 3240
tactggtaga ggtactttga tttggggagc tcgtactctg gataaaacgg ataactggcg 3300
ctatgttcct gtacgtcgtt tattcaatag cgcagagcga gatattaagt tagcaatgcg 3360
ttttgcagtt tttgagccta actcccaacc aatttgggaa aaggtcaagg ctgctatcaa 3420
tagctatttg cagtcacttt ggcagcaagg tgcactgcaa ggcaataaac ccgatgaagc 3480
ctggtttgta caaattggta aaggcgtgac catgacagat gatgatatta agaatgggag 3540
aatgattatc aaaatcggca tggcggcagt acgtccggca gaattcatta ttttacagtt 3600
tacgcagaat atcgcccagt aacttaggtc tataccctat agatttcaag atgcatcgcg 3660
gcggcaaggg agcgaatccc cgggagcata tacccaatag atttcaagtt gcagtgcggc 3720
ggcaagtgaa cgcatcccca ggagcataga taactatgtg actggggtaa gtgaacgcag 3780
ccaacaaagc agcagcttga aagatgaagg gtatagataa cgatgtgacc ggggtgagtg 3840
agtgcagcca acaaagaggc aacttgaaag ataacgggta tatttaatat gggcgattta 3900
ttgcccattt ttgtgaaagg aaatgagtta tgtcgccaac gctacccggt gtaacgatga 3960
ctcaggcgca gataacagcg ttcggtgtca gtacattaaa tatgcccgta ttcatagggt 4020
attgtacgag attgcctgcc ttttcagcgc ctgtaaaagt aaacagttta gctgaaacag 4080
aacaaataat agggaaagaa gggcgtttgt atgctctatt gcgccacttt ttcgataacg 4140
atgggataca agcttttatt ctgtcgttag gcgcacctgc tggggaaaat gctaatagtt 4200
ggcttgaggc attacaacag cccgatttgt atgcggctgt tgcagcagag ccgctaatta 4260
cacttttagc cgtcgttgag gcaagtgaac tgaaccaaaa agaaggtaat gaggctgtgg 4320
aagcttggcg acagtactgg aaagcagtat tagcgttatg tcaggcacgc agtgacttgt 4380
ttgccatatt ggaggcacca gatgataccg cattaatcaa gcgtagtttg caggattttc 4440
atcataaggc acgtcagttt ggcgctctct actggccaag gctagaaaca tcttatcaat 4500
cctctcagtt aaaaattttg tctcctattg gtgcagtagc agcggttatt caaagtaatg 4560
atgtccggcg aggggtagga catgcacctg ccaatatagc gttaaaacag acgattcgcc 4620
cgataaagtc ccgcctggaa ttagaagagt tgtatgaaga atcggatggt tcactgaatc 4680
tgatttgtag ttttccagct cgtggtactc gtatttgggg atgtcgtacg ttggcgggta 4740
ttgattcacc ttggcgttat attcaaaccc gattattgac ttcacacgtg gaaaggcaac 4800
tcagccagtt agggtgcatg ttgatgtttg aacctaataa cgcagtcact tggatgaagt 4860
ttaaaggcca tgctgggaat ctattaaggc agctttggtt acaaggggtg ctgtatgggc 4920
agcgtgaaga tgaagccttt tccgttgaaa tagatgaaaa cgaaacgatg actcgccagg 4980
atattgatga aggcagaatg attgctcgta ttcatttggc attgttagca ccggcagagt 5040
ttatcgctgt gactttgaat tttgatactc gctcaggcat tgcgacgagt acataataaa 5100
tcggaatatc tccatgacac taccagcaga gctttatacc ccagcggttt cacatcgttt 5160
tattgttaat tttcttttta aaggtttact tccttctccc gtagatattc gatttcaacg 5220
tgtttctggt ttagggcgtg agttacaggt tgaacagcgc catcaggggg gagaaaacgc 5280
acggaatcat tggttggctg aacgtataca gcataatagc ttgatattag aaagaggggt 5340
tatggtcgtt acccctttaa cactgatgtt tgatcaggtg atgcgggggg aaactctcaa 5400
ttgggcagat gtggtaatta ttcttctcga tcaggctcaa cgtccgataa caagttggac 5460
cttgagtcat gcgctaccgg ttcgctggca aacaggagat ttagatgcca acagtaacca 5520
agtgctgatt aacaccttag agctgcgtta tgaagatatg cgcattatag gggtaaaatt 5580
atgactatcg aaatccgtga actcattgtt caagcccgtg ttgtcgggac tgataccaaa 5640
acaacacgaa ccgttccttt atctattgtg caaatggaaa cacttataga acaacgtctg 5700
gttgaaaaag tgaagcggga gatattagac gtactccggg aagaacaagg tggtgggtta 5760
tgagcttgct tgaacgaggt ctggctaaac tcacgattac gggttggaag gagcgtgagc 5820
gtaaacatca gattggtaaa ctagaagcaa tgtataaccc ggaaacactt caactggatt 5880
atcaaactga ttatctccct gatgttagca ataatcaggt aacagtgagt aaccgctacg 5940
ttttgtcaaa gcccgcaggg ttaacactat ccttgttatt tgatgccaat atggctggtc 6000
ttacgacaac cgtcgagtcc caaatcacta ccctcaaatc gctttgttta gttaatgcaa 6060
gtactgatga acccaatttt ttggaaatta attggggggc aatgcgttgg gaaaataaaa 6120
attattttgt tggtcgggct agtggattgt ctctgactta tttgcgcttt gatcgtaacg 6180
caacaccatt gcgtgtgagt gcgcagctca cattagtcgc agatgaaagc tttgtgctcc 6240
aggataacca agccaagtta gatgcgccgc cggtatcagt agttaatgtc ccggatctga 6300
cttcattacc tgcactggcg aatatcgcta gcgtaaccac tatgttggga gtggattatt 6360
taatgttagc ccgcaccaat gatatggata atttggatga tatgcagcca ggtcagacat 6420
tgcgaacacc ggaggcatca tgagtttttt agataacagt aacttcaagc catcagatat 6480
caaactgttc gttaacattc agggagtgga gaaggaactc aacgaactga tagtaagcga 6540
attgaaaatc tcccgacgta tcaatgccat tccgcaggca gttgtaaagc taagagcgaa 6600
agagagtgaa agtggtgtat atcagtctga tgtacagcgg atgttgaaga gttgccgtcc 6660
gggagtaaag gcagagcttc gtattttgaa tacccggcta ttcagtggcg atattgtgca 6720
gcaaaaaaca gagttagtgt atgcgaaaac acacactatc aaattggtgc tacgccatga 6780
cttacagcgc atcaccggta attttcgtac cagagtgttt gcgaataccc gtgatcgtaa 6840
agtgatagcc gatctattga ataccgcaac attaaagccg gcattttcgg ggacatcaca 6900
ttgggatata gatcatgagc aactggttca gtatcgttgc agtgattggc aatttttgtt 6960
gcaacggctc tatgctacga atagctggtt gttagctgaa gaagataaag ataacactca 7020
ggggaaagtg accattattg ctccaaattc tttgcccctg aatgagcgtt ggacactgca 7080
acatcaggct gatcatcagg ctatccggct ttacagcacg gagctgatgc tggataaccg 7140
gtttgataca gcggaggctg ttgttagtgc ttgggatatt gatgatcagg cattactcgt 7200
ggcgtggaaa gaaaccctta gtcaagttgg gaaagatgcg ttagcgtcag ataattttag 7260
ccagacaaat aaagattcga gtgaactgtt attaagttgt ccgctctcta caaaagaagt 7320
tcaattttta acgcgtagcc aattagtcat gcggcgcttg acggccgttc gtggttcact 7380
gaaggttgaa ggcagtacta agtaccgttt agggcatgaa ctgatgttgt caggttttgg 7440
tgaaaatatg gatggctcac aaatactgac gggagtggat catcgaataa cggcagaaga 7500
aagttggaaa acaaccttac atgtgggatt agaactgccg ttaaaggcag agtatgtcac 7560
tcaggttaac ggtgttcata tcggcaaggt tgctgattat caatcagata gcaaaaaatg 7620
ggatcgtatt cctgttttga tccctgcatt tggaacgaat attcccttgt ttgcccgatt 7680
gggaaaaccc tacgccagcc accaaagtgg attttgtttc tatcctgaaa cgggtgatga 7740
agtcattctc agttttttgg aaggggaccc tcgttatcct gtcattattg attccctgca 7800
taatcctaaa caacagactc cattgcaaat cagcaaagag aataatctca aaatgttgat 7860
gattaagcag agcgataaag atgagcaaca attgttattt gatagccagc aacaaacagt 7920
cgcgttaatc ggtaagaaaa atatcgaggt taaaggtgag tatatcaacc tgactaaatc 7980
aaaggggact cgataatggc aaatacgctt attggccagg tatatggtca aggatgggct 8040
tttcccatta aatttattcc tgataataaa gaaaccgcag atcaaacagc cggtattgtt 8100
atggctcaag ggattgaaga tgtcagtcaa tcgctggaaa tattatttct taccgagcct 8160
ggcgaacgaa ttatgcgtga agattttggt tgtggtttac aagattttgt ttttgaaaat 8220
attagtgata cgctaatttc tgccatcaaa aatcgtattc agcaagcaat attacgttat 8280
gaacctcgcg catatttatt gaacgttgat attcaaacca aagaaaacca acctggacat 8340
ctgctcattc agattaattg gaaattacgt ggtagtgata tatctcagcg tttagacgga 8400
gtgcttagac tccattcagg tcaagcattg gaactgttat gaccaattat attattatcg 8460
acggggatct cattcaaata aatcccaaat ttgagggtga tcgaactctt acgattaatg 8520
gtattcctaa aataagcggg aatggagatg cgcaaattga aggaaaaaat atttgtgtgt 8580
caggtgatca cttaactgtc tcaattccag ccatttatat aacctccaga catcctgttg 8640
caggtagtgg aaaagtgaaa attacaaatt tatctgacga ccaactagca gaattttgtg 8700
ttagtgggga tgttgtgatt attgaaggca gtcagtttga agctcagttt acaccggata 8760
agccggccac taatccaagt aaccaagatg cagataatcc tgcgccttcg aatgggagtg 8820
ggagatttat acactcacag aacttcgtta aggcagaaaa ataaaaaatt ttgccgaagc 8880
ggttaataag tatgaataag cggggcggat aaaaacatgg atcttgctga attaaataat 8940
acgttgatga atgacttacc aacgaccaat tttaagttag aaacaaagga cccattaacg 9000
caattaaagt ggttacaacg ttatacagaa aatattcgtt tttatgcgaa tgatgattat 9060
ttctggcatc aattctggtt cttaaaaaat cacacaccag aagcgctctt tgctcgtttg 9120
caaggtgaaa cgttggctga tggagaattg cctcctcatc aagcgctatt gctggccttt 9180
ttacaacagc ttaagacgcc aggaatcatg cttgatactt tttcagcccg tcatcggcaa 9240
ttgtactatc aggaattgct agggataacg cagaaagatg cacaacctga tcatgtggcg 9300
cttggcgtgg tattaagtac tggtattgca gaatatttat taccgacagg cacattagtg 9360
gatggtggac aagacagcag cggaaattca ctgcaatatg cgttggatac cgatttattg 9420
gttaatccag ggcaattaac agatgttcgc tacagctatt tggatcataa gacctataaa 9480
atcttcatct tgcaagatga taaagcgaat atcagttggc cctcttcagg cgctcgttta 9540
tttgtagcac ctgagggcaa cggacaggaa aaggcacctg aacaaaagtt ggcactttac 9600
ctgggatttg atgatataca gccagggcaa actctttctt tattttggca attcattgca 9660
tcaactcccc tgacattaaa atggttttat ctgaacgaga taaataactg ggtgaagcta 9720
gatagtgtca gagataacac ggatggcttt tttatcagtg gattatggca agcgatatta 9780
cctgatgatg cggtgaaaat gtattttcca gagacaactt ctgtaaaacg ctactggatt 9840
aaagctgagg tggaatcgct tactgaatct ggcgatttgt ggcaaccgct attagaaggc 9900
atcttgtata acgctcaaac agcaacgctg gttgatgcag acaacacaga tgaaaagcac 9960
tttcatgatg ggctgatgcc ttttagcgtg cagcatttgg tcaacaccgt ttcagaggta 10020
aaaaaaattg agcagccctg gtcttcttgg gggggaacgc cacaggaaga cactactgat 10080
ttcttccatc gagcggcaac acgtcttcag catcgccagc gtgcgttaac ttgggataac 10140
caaattgcca tgttgaaggc tgaatttccg cggatttatg atgtcatctc accaaatatc 10200
acgtggatga accaacttca gacatcaaat acgcaaacgc tgatcgttat tcctgatgtg 10260
aactacagcg acaacaagga tcgcttacgg ccacaattca gccctgccag cttgcgacaa 10320
atgagtgact ggttacagat tcacactagc gcatgggcga atccacaagt ggaaaatcca 10380
atttatattg atgtctctgt gacctatgag gtgcaattta gtgcgggtgt gaatcctgat 10440
tatgccctcc ggcaattaca acaatggttg agttcaattt atatgccatg gtatcacgca 10500
gataaaaaag gtgttgccgc tggcgatcaa atcgattttt accaactgtt tgcagatatt 10560
cagcgagtac cttacgtgga gcatgtcaaa acattgacat tgaccacaaa agacacctca 10620
ttaaccaatg gcggggttat taaggcacag caaaatgaag tgctggtgtt ggtatggcaa 10680
caaggagaac aaattaggca gggagaatcg aaatgaggca gcataatgag ttatttcctg 10740
tagtaaaaga cgcgataagc tttgaaaacc tgcaagctca gggtgagaag gttattagtg 10800
atcagtccgg taacatatgg agcgataaag ataaacatga tcctggtata acattactag 10860
actctttaag ttacggtgtt tcggatttag cgtatcggca ctcattacct ttaaccgatt 10920
tattaaccat tgctggaaaa gatacgcttt ttccagccga attcgggcca cagcagacgc 10980
taacttgtgg ccctataaca ctggatgatt accggcgtgc gttacttgat ttacatggta 11040
atgatgcatt taaaatatca gctagtgacc ccagagactt tttgtttcag gatatacagt 11100
taatttgtga gccaaaaagt aagcgttata aatactattt caatcccgaa acgcttgaat 11160
atacattcac gccaccttca ggggataaat ttaaaacttt aacactacga gggaattatt 11220
ggctttattg gataccaacc cgttgggcag gtaaatcagc taatttgccg ttagttaagc 11280
gggtgatgga agattttctc cgtgaaaatc gaaatttggg ggaaaatgtt gttcaagtga 11340
cacgggtgat atcaacgcct atttatcctg agctggtcat tgagctggcg gatgatatta 11400
cagatgcggc atcagtatta gcatcaatct atatgctatt agaacagtgg gcgatgccga 11460
tgcctgctcg ctttactacc gaagcattac aggccaaggg attaacaaac gaagagatct 11520
ttgatgggcc gtggttgcgt catggttgga tacctcagtt accgacctct caaaactacc 11580
atacaggcat ggttctgaag atgaatcatc tgattaacca attgctggcg gttgaaggta 11640
taaagcgcgt agttagcctg acgttgccag aaacagaata tttgcatcag ataaaagatg 11700
ataattggtc ctggcaatta gatgttggtt attatccatt attatgggga gctaatccac 11760
tagaggtaat tacagagaaa aataacaatt atgtcaaatt gttcgcaaaa ggtggggtac 11820
gattacaacc tgatcagaaa agtgttgagc ggttattatc acaggaatca ctcattaata 11880
atgctgcatc cacgttaccg gctggtaagg tgcgtgatct caaagcctat acacctataa 11940
gccgcaggtt gcctgcctgt tatggtttgc agaatacttt gcaaaagtta aaacctgaac 12000
aacgacactt atatcagttc ctattaccat tggagcaaat gcttgctgat ggatgtgcgc 12060
ggcttgcatt tttgccacat ttgttagcat ttagggaccg aagcggaaat atcagtgata 12120
cactctggcc tttcaagaat acagaggaca caattgccca acaggttcat caggaatatg 12180
ccggtacatt aaaagccttt caacagcagg aaattagcct gtttgatgat aaaaatagac 12240
cgcatcatgg caatatcaat cgggaattag atattcttga ttatctgcta gggtattttg 12300
gtacacaacg tgcaaagcgt ccattaacgc aggatattca tgattttctg caaacccagc 12360
gaggttattt ggcacagcag ccggagttgg gttatcagcg tgataatatc cgtattgatc 12420
gagtttcagc tttacaaaaa cgtatagcag cccgaattgg gctagatggt actattttca 12480
aagaatcggt tgatttaagt aagttacctt tttatttgat tgaacatcgt cagcttttac 12540
caaatttacc ccatcttgac tttcaacatg atcaaactcc ccaatctttt gtgatttccg 12600
acaacattgt taaagtgaaa caagcgggaa tagcagataa aatcgttcgt ggacagctta 12660
ttgattttat agatattgaa agcaaattta ccgttcgtgc ccaaatgatt gtcgctgtag 12720
agggaaatga attttctctg gatacaaaaa atagtattca acttgaaaag aatctgcagt 12780
tattacaatc agcgtctgag aaaaacaatt tacgatggag aaatagcacg gcgtggttag 12840
aggatatgac gtatcgtatc aattatactg acgatcaggt tatagacgat aaaacaaaac 12900
aatgtcgttt acaaagtaat actaaatcgc cttttccagc cttaattgca ccaaaaaata 12960
agattacgat tattaagcaa tcttctccac tctccagtat tgctgaattt actgatgaac 13020
cagaattcaa attagttgca acggtgacag agattgatcg gattgaaggg atattgacta 13080
tcgaacggga tgacaaccaa ctccctttcc cgactaaaga agagagtaat caatatatat 13140
ggtacatatc tgatgaaaac tatatttcaa gtgatcgttt ctcttttgtg gtgagcgtcg 13200
tgctgaatcg cggtttggtt gaaagggaag atattgatca atataagcta gaggaatgga 13260
tagagcgtga aacacttgca gagtttcctg cacatatttc gttaattact cattggctgg 13320
catctgaaaa tttcgatgat tttgcgaaga catatcaacg ttggcaaaac aatggggcgc 13380
agttagggga tgaatcctac accattttgg aaaaactgac attagggcat ttaccaacag 13440
gacttactgg cattagtaat atgtttattg ctacagaagc tcagcgtcta gaagttgttg 13500
gcgagagtgg taatgagtgg aatacccagg caattattaa caacgaacta ttctatgttc 13560
cctcacagaa tagttaatac cgagtgttgt gatcaacttt tattataagc cggaggataa 13620
atggacaaca aaaataacaa acctactgat caagagattc taaaaacatc acgggctgtc 13680
ggagaaattc cttcagcgga taatttaaaa aatcgtttta aagctcgttc gattccatta 13740
gagacggatt ttactaatct cattgacctt gctgaagttg gacgattggc tatcggccag 13800
tcaccatcgc agcaaagtaa aacgcctggc accggaatgg aattaacttc ggatggtaaa 13860
ttacaagtca aggctggggc aggtgttgat atcgataata ataatcgtat tactattaag 13920
tctggtcatg gaattaaggt tgatggaaac ggcatttccg ttaaaccagg ttcgggtatt 13980
aaggttgata gtaatggtgt aaatgtcaat attgatgatt tttgggagga aatacgcaat 14040
aaaattatgc ctaaaggaac catgctgcct atttatggca cacctaaccc ctctgcgctg 14100
ccaacaggat gggaatggtg tgatggtaaa gatggcagac ctaatttaaa aaaagggaaa 14160
tataacttac tatcaggtca gtcttcaggt actgatactt tttgggcaga taataagaat 14220
ggagatacag agatcaacgt gttatttgtt tactatatga ttaaggttgt gtaatatctt 14280
aagtaatatg cattactcta aaatgaatga tttatattta agtaacataa taattaagtt 14340
gtgttgtagg gctgttttta tgagaaatat aaaaacggag gtaataattg gcttcaaaat 14400
atcagtgatg aaatagagtt atttcgcttt ataaaaattt tgttttattt cttttaataa 14460
ttatttatag aaggtaatga tatgtgcaca caaaaaaacg tgttagatag actgaaagat 14520
agaaatatta cattgggttg ggatgttgtt gttgcatata accaagaaag tgttaataag 14580
ttattgaagc aacaatatgt tgaaaaagtt tactcaaatg aacattttgt ttttaaagat 14640
tggcatgatg ataataaaac gaaatttatt gagggattaa cagtaggcgc tccactagtt 14700
tcatttgagg aggcgtcttt atccgatgct aatgtaaaag tgacacttaa ctttctttct 14760
ggtagatgga gagttataca agcaaatacc ggcacaccaa ttgaatggaa agaaattgtt 14820
cctggcagtg gctataaagc agaattagtt gttccgctta aatcaataac tggtagtgta 14880
agtaaaaaag atatcatatt aaaattcaaa gatgctgtcg taaaaaaaat aaatttattt 14940
gacaatcaag agcctgattt tattaattat ttcaagcaat cgatcagtga gggaaattat 15000
actttagggc aactggtgac agacagcaca ccgggattaa ttcctgctga atttcatatt 15060
cgtactcaac cccatccaaa aacacgtgag cgtggttctc aatatgtagg aaatggtgcg 15120
gtactgttgt ttattaaaac gcaatatggc ggaagtggaa cattgcctgt aaatgatttt 15180
gattggttaa ttcctgatga tcatactagc gcattagtca tttcgagtaa gaccatgatg 15240
gggcaaatat tgccaaaaca atacaaagat aaattgcctg gtgatcctca gtttagccca 15300
ccaaaaagag tcaatgataa acaagactct gcttattata ttacgattac cgatggtgga 15360
tttgatggta atagccctat agagaagtca tggttacgtt ctgattatag caatgggatt 15420
tggactggtg aacgtggtaa tgctattatt ggtgaaaaag gaaagcggat accaccacgt 15480
tttccatacc aaaattttgt tattaaacct catggtgaat cgttatttca aggatgggag 15540
aataagataa attacactca aaagtgtgca agatatttcc gacatcatag taatagtata 15600
actttcgaag atactgcatt aatggatctc agtattggtg gacaaggtag tatcaattgc 15660
cagattgatg gtgaacattt ctatttaaaa tcagatgatt tttcccccaa tgtcagctat 15720
gaaccaactt cattctggga taaatttatc ggtggggtgg atgcaaatgt gaaagatgaa 15780
ttcagagatg aattagcaca acaggcagaa gcaaagttaa aacaggtatt taatattgaa 15840
ttgcctgaaa tcagtctgtt ttctattaaa catctgctct ttcctggcat ggatgttatg 15900
caacttaaac agggttattt cccaggagat ttgattatct ttggggatat ttcacctaaa 15960
ttgaccacaa ttcaggtggc tcctttggaa gccatggttg cccttaaaga aaatcaaaaa 16020
ttcactgtcg tacctgaaaa taaaaatgtt agttggaagt tggatcataa tagtgaggct 16080
atcaatgatc cgggaaatat tgatgataaa ggtatttata cggcaccggg cagaatcaga 16140
tctggttctg aagtcattaa agtcactgca actgacggcg atggaaatca ggcatcggcg 16200
gcgctgacgt tggttccttc ttctgttgca ttaacacctt cttttgcttt tatctctgaa 16260
gcagataaga aacctatatt attattggcg aatgtcctag acggaaaagc agtaacatgg 16320
aatgtggaaa gctgtacagg cagccaatgt ggttctgttg atcagaatgg gctttatact 16380
ccaccagcag ggcgttttaa cgatggattt acttttgcat ccatcaccgc aactgcaaaa 16440
gatggtagtc aagcacgaac cattatttgt ctaatggcat caatgccagg acatggtttt 16500
tacaaggttg aacctaattt acgtttgaat gtgaaagtag gggaagaaat tatctttaaa 16560
gcgcaggcag atagctataa tggtgatcct gatacttggg aaattttccc tcctcgcgga 16620
aaattaagtg aacctgagtt tgaacccaat aatgatcctg aaactaatga tacaattttt 16680
ggtcattata aggtgaccta taccgcgccg actaatgtta cctcacctga attgcttgtt 16740
gtccatgtat gggagaaaaa taggcataat gagaaaaaca aaggtaaggc aggatatgca 16800
cttattgaaa ttatcccaga tgataaatag aaaatttatt taaataaaaa tcacagcggg 16860
tttatctcgc tgtgattaaa gtcatctttt tttatagatt gtttatctct aataataatt 16920
ttattttata atataaagga aattaaaatg aataatgaat ataaaaataa caccgtgaat 16980
tggcgtattt cacctgatac ggtaggaagt attgataata acggtttata tacagcacct 17040
aatcgggtaa agaatatcga atttgtccaa gtaatggcaa gcgatgctaa taataatcaa 17100
tcttctgcga ttattactgt tattccctct tctgttgcgt taacgccatc gtttactttt 17160
atctctgagg caaaaaaaac atcagtcact tttaaagcga cagaacttga agggaaaaaa 17220
gtgacatgga gtataaataa ttataccagt aatcagtatg gttccatcga tcaaaatggt 17280
atctacacac caccggaaag tcgttttaac gatggatata cttttgtatc tattacagca 17340
aaagcggaaa atggcgctga agcgcaagcg cttatttgct tgatggccaa aattccaggg 17400
catgcctttt tcgatgttca gcctaatata tgtttaagtg tgaagcctgg agaagaaatc 17460
atttttagag ctaacgcaga tcgttataat ggtgatcctg attcctggga aattttcccg 17520
tctcttggta aattgggtga gcctgagtat ataaaaaata acgatccaga aattcctatt 17580
tatggatatt atcaagtgaa atatattgcg ccaaccaata taaattcttc ccaaatactc 17640
gttgtgcgta cttgggaata tgacaaacat gatgagcata atcaaggtaa agcaggatat 17700
gcattcattg aaattgtgcc agaaaatgag ctttaatata tatacccaat agatttcgag 17760
ccgcagcgcg gcggcaagtg aatgaatccc caggagcata gataacgatg tgactggggt 17820
gagtgaacgc agccaacaaa gaggtaactt gaaagataat gagtataaat gactttagta 17880
agagaaatta tggcttcatt cagaactatt tattagagta attaacttta taaagacatt 17940
taatggaaaa tataatagaa aaatttaata ttaatattga agtctcatct gaaattattg 18000
gagagagttt attaaactcc cctttattga tgagtagaga aatcagcaat caattatctg 18060
aaatattatt agattataaa gaatataata ttgcattgga taagttagtg ttaaatatag 18120
gagaaatacc ctatgaaata tttgaacaac aattctatgg tcgtttggga aaattattaa 18180
atgaaaagtt aacaataata ataaatgata aattattggt aaaaaacata tcaacctcgt 18240
tatttcctga atgttttagt gaaaaaagaa acccattatt aaatagagtc ataaaaaatt 18300
taccttctaa tttggttttt gaagttcatt caatggtaaa aatagaatca gtaaataaca 18360
aaaaacaagc taatatattg acatcttatc tggcttattc tttttttaat aaaagcaaat 18420
tacaacaaca tttattttcc actagtaata ataaattaat tgagagctta tacgcacttt 18480
ttctaacgga tcagaatcga atacctactg ctcataaaat aggaaaaggt gcacttatac 18540
tatctgccct tatttggctt tattctaatt ccaatgatta tctgcccaaa ccagaaagca 18600
ctctgttgtt acaaatagaa caggatataa aacaaggata tttgccttta acgttgttaa 18660
tcactttctt ccagaacaga aatggcgggc gtgttttttg cgattggcag tatgcgttat 18720
ggcaaatcga tatcatcaaa aatcacttag gcattaaaat aacatcgaaa gaaccccatt 18780
tacgggagaa aataatgtta caaccagtta atgcttctga tcgatcctct gtgctgatat 18840
cagacgaaaa attgacaata ccgttaacaa ttacaggtgc gggattagtg cttctctggc 18900
cactattaac tccactattt tcgtcttttg atttgttaga taagaaaagt ttttcagaca 18960
atttggcaca ggaaatagca tttaatttat tggaatggtt agtctgggga gatgagatgc 19020
tgttacatca ggaatcatca ttatctttat tactctgcgg aatagatcac caaacaatac 19080
tggagcgcca ggttcttatt cctgagcaca aggaaaaatt aaataactgg ttgcaaggta 19140
tttgtactca acttttctct tggaaaaagc tagggatcga tgatatgcgc caactttttt 19200
tgcagcgtca ggctgcactt tattatgaag atgatggccg ttggttatta acggtgcagc 19260
gtgaagctta tgatgtatta ctgactcaaa tgccttggcc gtggccattg aatattgtga 19320
cattaccttg gctagctgag ccgattagta tcacttggga aggtatctct gaaccaacgg 19380
atttgtcatt ttggtaatcc aatatctcat taggaactct atgcatgtac gatttatctg 19440
atgatcttgc cagacagaat atttcaccgg aatatgaatt gacggttttg ctgtctcaga 19500
ctgctatatt ggataaacga attcgtttac gaattcagga attaatgcaa cagcaaacac 19560
tattgggaga aagtggacag acgtcttttg atgatatttc attttcattc gtttcgagtg 19620
aacaacaaaa atcatcttat ttggtgtcac cgcatcaaaa ttggacgaaa gaggattttc 19680
ctcctgagcc gatcccatct cgtagccgtc taggacaatt agttgaacgg tttgacttaa 19740
ctcaatttga aattgatttg attttattgt gcctgttgcc tcatcttgac agacgttatc 19800
taacgttatt ttctcttgtt ccggtaagtg gaggtaataa cagcaaaaag cagatgttaa 19860
cgttgggatt ggctttggag ttgctttgtc cgagtgtagt agagcgcaat gcgcaacgtg 19920
ccagtttatt accacaggca ccgctttggg attatcgttt atttcagttg cgcggtgata 19980
tgtctgtttc ctacgatgaa ataccgttag caatcgataa ttctcttatg cattggttat 20040
tggggcatga tgctctcccg atttctcttc tctcccgggc tcattggctt cctgttcctg 20100
aagtgcctga tattttgcct gatttcacca accaattgat agaactctgc caaatggaac 20160
aagaggggat gctgacaata atcgccggcg gagccggaag tggcagcaaa acaagtgttg 20220
cacgcgcagc atcacaagta gggcgctctg tattgttgtt atcgttagca tcagtgacac 20280
tgagtgaaca tgaaactatt acactgataa cactggcatt acgtgaagca caactaagaa 20340
atgcctgtct tatgtttgaa gctttggatg agttttgtga agcacgcccc gctttgcagc 20400
tctggctagg aaatcgactg gctcgttgtt cgattccgct gttttgtcaa ttacctaagc 20460
aagcatcatt attgccattg gatgcaattt cacaagttgt attgtctatg ccaatgcctt 20520
ctttaatggt gaaggctgca gcattagctt caatgatgac gaattatttt ccagacaatt 20580
cattggatgt tgaaagttta gtgacatgtt tccatccttc tccattgata ttgaaaaagg 20640
cccttagtga agcagaaatt tatcgccgac tacgggggga aacggctagt ttgagattag 20700
atgatgtgca aatgtccctg cgttttcggt tacagcagaa ttttggacgt ttagcacaga 20760
gaattacacc acaacgaacc tttgatgatt tgatcatcag tgaatctcaa cagcaacaat 20820
tacaagaaat cctggcggct attcggcaac gagataggat gctagagcaa ggatttgctc 20880
gtaaagtgag ctatgggacg ggtatcagca cgctattttt tggtgaatct ggcacaggaa 20940
aaacgatggt agcagaagtg ttagctggtg ttttaggtgt ggatttgatc aaggtagatt 21000
tgtccactgt ggttaacaaa tatattggtg aaactgaaaa aaatctggct cgtgtttttg 21060
attatgccca agaagacgcc ggggtattgt tctttgatga ggcagatgca ttgtttggca 21120
aacgaagtga aactaaagat gcaaaagatc gtcatgctaa tattgaagtt tcctacctat 21180
tgcaacgcct tgaaagttat ccagggctgg tgatattagc caccaattac cgtaatcatt 21240
tagactcagc atttagtcgt cgcctgactt tttcggtacg attctctttt ccagatgttt 21300
ccttacggga acggatgtgg cggattatct ggccatcggg aattcaatta gccgacgaca 21360
tcagtttttc agcgttggca aaacgggctg aattaacggg ggcgaatatc cgtaatattg 21420
cgctactcgc tagttggctg gcagtagatg aaggaaatga aaaaattact atggctcata 21480
ttgaatgcgc attacgacgt gaactgagta aagttgggcg cattgattta ccttaatttt 21540
tctttgtaat cgggagacaa ctatggttaa aaatatcaaa tcagatgaaa ccttactgat 21600
attaaatagt aaaatagaag atgcattaaa agcgtattta ccgggcgaag atgtcgttat 21660
tcggttcgat atgtttggta aaaatgaaaa tccagattct cctaccgtgt gcgtttttct 21720
ttatgatatt caggaagatc tgcaattacg cgtgggagaa gggcggcaat acctgcctgc 21780
gacaggaaat tttgtcccgg gatgtgtcaa tgttcgttgt aattatctta tttcctactg 21840
ggagccggaa cagagcggag ggcagggatc gccaaccata cgttctaata gtcaatcaat 21900
gaagataatg aactgtgtat tgaatgcatt aattaatcat cgttcatttc ctggtttacc 21960
cagaacttat acgagagttc ttcctcctaa tgaacaatta aatagcttag gaaacttttg 22020
gcaatcatta gataataagc ctcgactatg tttaagttat atggtgacta ttcctattca 22080
acttaccccg ccgacagaga aggtatctcc tgtcattacc tcaaaaactg atattactcg 22140
aaaaccatcg cttaactttt atcttgaggc agatgaaatt atccgtcagg cattagttga 22200
tgccttaata tctcaaacaa cagaatctat ggatacgata actagctggc tggcaaaagt 22260
tgttattatt tgtcgaccac cagaaataat gaataaacaa atgattgaac aaactgtgaa 22320
attaattatt gctggaatta cagaagaggg attagctgga aatataaaga caatcactca 22380
aaagtgggtg gaagagaaga cgattattgg tgaaatcgac gatgtttctc tagttatttc 22440
ccaagttgac acgacagcgt tgtctgctgt aacaataccg acatctgttt aa 22492
<210> 96
<211> 14
<212> PRT
<213> 光杆状菌(Photorhabdus)
<400> 96
Thr Gly Gln Lys Pro Gly Asn Asn Glu Trp Lys Thr Gly Arg
1 5 10
<210> 97
<211> 34
<212> DNA
<213> 人工序列(Artificial Sequence)
<220>
<221> primer_bind
<222> (1)..(34)
<223> 引物
<400> 97
tatcatatgt ctacaactcc agaacaaatt gctg 34
<210> 98
<211> 27
<212> DNA
<213> 人工序列(Artificial Sequence)
<220>
<221> unsure
<222> (1)..(27)
<223> 人工
<400> 98
atctctagaa cagatattcc agccagc 27
<210> 99
<211> 19
<212> DNA
<213> 人工序列(Artificial Sequence)
<220>
<221> primer_bind
<222> (1)..(19)
<223> 引物
<400> 99
ggcgtcacac tttgctatg 19
<210> 100
<211> 20
<212> DNA
<213> 人工序列(Artificial Sequence)
<220>
<221> primer_bind
<222> (1)..(20)
<223> 引物
<400> 100
tcggtggcag taaattgtcc 20
<210> 101
<211> 27
<212> DNA
<213> 人工序列(Artificial Sequence)
<220>
<221> primer_bind
<222> (1)..(27)
<223> 引物
<400> 101
atgtctacaa gtacatctca aattgcg 27
<210> 102
<211> 19
<212> DNA
<213> 人工序列(Artificial Sequence)
<220>
<221> primer_bind
<222> (1)..(19)
<223> 引物
<400> 102
gactcccttg agggtacgg 19
<210> 103
<211> 21
<212> DNA
<213> 人工序列(Artificial Sequence)
<220>
<221> primer_bind
<222> (1)..(21)
<223> 引物
<400> 103
ttctgatgag agtgatggta c 21
<210> 104
<211> 24
<212> DNA
<213> 人工序列(Artificial Sequence)
<220>
<221> primer_bind
<222> (1)..(24)
<223> 引物
<400> 104
tgaataaaga attcagtcaa tatc 24
<210> 105
<211> 20
<212> DNA
<213> 人工序列(Artificial Sequence)
<220>
<221> primer_bind
<222> (1)..(20)
<223> 引物
<400> 105
tagtggctga tgaaagtctg 20
<210> 106
<211> 22
<212> DNA
<213> 人工序列(Artificial Sequence)
<220>
<221> primer_bind
<222> (1)..(22)
<223> 引物
<400> 106
ggaagccaaa gataatgaag tg 22
<210> 107
<211> 19
<212> DNA
<213> 人工序列(Artificial Sequence)
<220>
<221> primer_bind
<222> (1)..(19)
<223> 引物
<400> 107
catttcttcc ctatggttg 19
<210> 108
<211> 24
<212> DNA
<213> 人工序列(Artificial Sequence)
<220>
<221> primer_bind
<222> (1)..(24)
<223> 引物
<400> 108
ttaaattcct acaagattat cttt 24
<210> 109
<211> 133
<212> PRT
<213> 智人(Homo sapiens)
<400> 109
Arg Ser Ser His Ser Arg Leu Gly Arg Ile Glu Ala Asp Ser Glu Ser
1 5 10 15
Gln Glu Asp Ile Ile Arg Asn Ile Ala Arg His Leu Ala Gln Val Gly
20 25 30
Asp Ser Met Asp Arg Ser Ile Pro Pro Gly Leu Val Asn Gly Leu Ala
35 40 45
Leu Gln Leu Arg Asn Thr Ser Arg Ser Glu Glu Asp Arg Asn Arg Asp
50 55 60
Leu Ala Thr Ala Leu Glu Gln Leu Leu Gln Ala Tyr Pro Arg Asp Met
65 70 75 80
Glu Lys Glu Lys Thr Met Leu Val Leu Ala Leu Leu Leu Ala Lys Lys
85 90 95
Val Ala Ser His Thr Pro Ser Leu Leu Arg Asp Val Phe His Thr Thr
100 105 110
Val Asn Phe Ile Asn Gln Asn Leu Arg Thr Tyr Val Arg Ser Leu Ala
115 120 125
Arg Asn Gly Met Asp
130
<210> 110
<211> 399
<212> DNA
<213> 人工序列(Artificial Sequence)
<220>
<221> primer_bind
<222> (1)..(399)
<223> 人工
<400> 110
cggtcaagtc actcgcgtct ggggagaatc gaggctgata gtgagagcca agaggatatc 60
ataagaaaca tagcacgcca tttggcacag gtaggcgatt ctatggatcg ctccatcccg 120
cctggacttg tcaatggtct tgcgcttcaa cttcgtaaca cttcccggtc cgaggaagac 180
agaaatcggg accttgcgac tgctctggaa caactgcttc aagcatatcc tcgtgacatg 240
gagaaagaaa agactatgtt agtattagct cttcttttag ctaaaaaggt agcttcgcac 300
actccaagtt tattgcggga cgtttttcac accactgtta atttcatcaa tcagaacctg 360
cgtacttatg tgagatcttt ggcgagaaat ggtatggat 399
<210> 111
<211> 15
<212> PRT
<213> 智人(Homo sapiens)
<400> 111
Leu Ser Glu Ser Leu Lys Arg Ile Gly Asp Glu Leu Asp Ser Asn
1 5 10 15
<210> 112
<211> 45
<212> DNA
<213> 人工序列(Artificial Sequence)
<220>
<221> misc_feature
<222> (1)..(45)
<223> 人工
<400> 112
ctgtcggaga gtttgaagcg tataggtgac gagctggaca gcaat 45
Claims (25)
1.光杆状菌毒力盒(PVC)效应子前导序列用于将有效载荷包装到PVC针状复合物中的用途；
其中有效载荷是选自多肽、核酸或其组合的一种或多种；和
其中前导序列和有效载荷形成不同于野生型PVC效应子蛋白的效应子融合体。
2.根据权利要求1的用途，其中所述前导序列包含PVC效应子的氨基酸残基1-50。
3.根据权利要求1或权利要求2的用途，其中前导序列包含与选自SEQ ID NO.:47-SEQID NO.:92的一个或多个序列具有至少60％序列同一性的氨基酸序列。
4.根据之前任一项权利要求中的用途，其中PVC效应子包含选自SEQ ID NO.:1-SEQ IDNO.:46的一个或多个序列的氨基酸序列。
5.根据之前任一项权利要求中的用途，其中PVC效应子包含选自SEQ ID NO:4、SEQ IDNO:22、SEQ ID NO:25、SEQ ID NO:30、SEQ ID NO:32和SEQ ID NO:46的序列。
6.根据之前任一项权利要求中的用途，其中前导序列与有效载荷共价融合，优选在有效载荷的N-末端。
7.一种用于制造包含有效载荷的PVC针状复合物的方法，该方法包括：
a.使PVC针状复合物与包含与有效载荷融合的PVC效应子前导序列的效应子融合体接触；
b.其中所述有效载荷是选自多肽、核酸或其组合的一种或多种；和
c.其中效应子融合体不同于野生型PVC效应子蛋白。
8.根据权利要求7的方法，其中所述接触在细胞内、细胞裂解物中或纯化的细胞裂解物中进行。
9.一种用于将有效载荷递送到细胞中的体外和/或离体方法，该方法包括：
a.使细胞接触包含效应子融合体的PVC针状复合物；
b.其中效应子融合体包含与有效载荷融合的PVC效应子前导序列；
c.其中所述有效载荷是选自多肽、核酸或其组合的一种或多种；
d.其中效应子融合体不同于野生型PVC效应子蛋白。
10.一种用于遏制害虫的方法，该方法包括：
a.使害虫或包含害虫的目标区域与包含效应子融合体的PVC针状复合物接触；
b.其中效应子融合体包含与有效载荷融合的PVC效应子前导序列；
c.其中所述有效载荷是选自多肽、核酸或其组合的一种或多种；
d.其中效应子融合体不同于野生型PVC效应子蛋白。
11.一种PVC针状复合物，用于处理方法中；
a.其中PVC针状复合物包含效应子融合体，该效应子融合体包含与有效载荷融合的PVC效应子前导序列；
b.其中所述有效载荷是选自多肽、核酸或其组合的一种或多种；和
c.其中效应子融合体不同于野生型PVC效应子蛋白。
12.一种包含效应子融合体的PVC针状复合物；
a.其中所述效应子融合体包含与有效载荷融合的PVC效应子前导序列；
b.其中所述有效载荷是选自多肽、核酸或其组合的一种或多种；和
c.其中效应子融合体不同于野生型PVC效应子蛋白。
13.一种效应子融合体，包含与有效载荷融合的PVC效应子前导序列；
a.其中所述有效载荷是选自多肽、核酸或其组合的一种或多种；和
b.其中效应子融合体不同于野生型PVC效应子蛋白。
14.一种分离的PVC效应子前导序列。
15.根据权利要求7-14任一项的方法、用于使用的PVC针状复合物、PVC针状复合物、效应子融合体，或分离的PVC效应子前导序列，其中前导序列包含PVC效应子的氨基酸残基1-50。
16.根据权利要求7-15任一项的方法、用于使用的PVC针状复合物、PVC针状复合物、效应子融合体，或分离的PVC效应子前导序列，其中前导序列包含与选自SEQ ID NO.:47-SEQID NO.:92的一个或多个序列具有至少60％序列同一性的氨基酸序列。
17.根据权利要求7-16任一项的方法、用于使用的PVC针状复合物、PVC针状复合物、效应子融合体，或分离的PVC效应子前导序列，其中PVC效应子包含选自SEQ ID NO.:1-SEQ IDNO.:46的一个或多个序列的氨基酸序列。
18.根据权利要求7-17任一项的方法、用于使用的PVC针状复合物、PVC针状复合物、效应子融合体，或分离的PVC效应子前导序列，其中PVC效应子包含选自SEQ ID NO:4、SEQ IDNO:22、SEQ ID NO:25、SEQ ID NO:30、SEQ ID NO:32和SEQ ID NO:46的序列。
19.根据权利要求7-18任一项的方法、用于使用的PVC针状复合物、PVC针状复合物、效应子融合体，或分离的PVC效应子前导序列，其中前导序列与有效载荷共价融合。
20.一种包含核苷酸序列的分离核酸，所述核苷酸序列编码权利要求14-19任一项的分离的PVC效应子前导序列。
21.一种包含权利要求20的分离的核酸分子的表达载体。
22.一种包含权利要求20的分离的核酸分子，或权利要求21的表达载体的宿主细胞。
23.权利要求22的宿主细胞，其中所述宿主细胞是选自哺乳动物细胞、昆虫细胞、酵母细胞、细菌细胞，和/或植物细胞的一种或多种；优选其中所述细菌细胞是大肠杆菌细胞。
24.权利要求22的宿主细胞，其中所述宿主细胞是光杆状菌属细胞。
25.权利要求24的宿主细胞，其中所述光杆状菌属细胞包含与诱导型启动子可操作地连接的光杆状菌PVC操纵子。
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| GBGB1908155.3A GB201908155D0 (en) | 2019-06-07 | 2019-06-07 | Leader sequence |
| GB1908155.3 | 2019-06-07 | ||
| PCT/GB2020/051380 WO2020245611A1 (en) | 2019-06-07 | 2020-06-05 | Leader sequence |
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| EP (1) | EP3980545A1 (zh) |
| JP (1) | JP2022535283A (zh) |
| KR (1) | KR20220133757A (zh) |
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| CA (1) | CA3142462A1 (zh) |
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| WO2024003387A1 (en) | 2022-06-30 | 2024-01-04 | Københavns Universitet | Contractile injection system and use thereof |
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| US20160046951A1 (en) * | 2013-03-07 | 2016-02-18 | The General Hospital Corporation | Compositions and methods for bacterial delivery of polypeptides |
| CN108137649A (zh) * | 2015-10-14 | 2018-06-08 | 拜耳作物科学有限合伙公司 | AXMI554δ-内毒素基因及其使用方法 |
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| US5223409A (en) | 1988-09-02 | 1993-06-29 | Protein Engineering Corp. | Directed evolution of novel binding proteins |
| IL99552A0 (en) | 1990-09-28 | 1992-08-18 | Ixsys Inc | Compositions containing procaryotic cells,a kit for the preparation of vectors useful for the coexpression of two or more dna sequences and methods for the use thereof |
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20160046951A1 (en) * | 2013-03-07 | 2016-02-18 | The General Hospital Corporation | Compositions and methods for bacterial delivery of polypeptides |
| CN108137649A (zh) * | 2015-10-14 | 2018-06-08 | 拜耳作物科学有限合伙公司 | AXMI554δ-内毒素基因及其使用方法 |
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| CA3142462A1 (en) | 2020-12-10 |
| AU2020288380A1 (en) | 2022-01-06 |
| GB201908155D0 (en) | 2019-07-24 |
| MX2021015121A (es) | 2022-03-17 |
| JP2022535283A (ja) | 2022-08-05 |
| SG11202113297RA (en) | 2021-12-30 |
| KR20220133757A (ko) | 2022-10-05 |
| WO2020245611A1 (en) | 2020-12-10 |
| BR112021024592A2 (pt) | 2022-01-18 |
| EP3980545A1 (en) | 2022-04-13 |
| US20230076614A1 (en) | 2023-03-09 |
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