CN111826398A - 用于活细胞间膜蛋白展示和相互作用检测的工程质粒系统 - Google Patents
用于活细胞间膜蛋白展示和相互作用检测的工程质粒系统 Download PDFInfo
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Abstract
本发明公开了一种用于活细胞间膜蛋白展示和相互作用检测的工程质粒系统。该工程质粒系统包含配体‑受体信号识别质粒和信号激活质粒,其中配体‑受体信号识别质粒包含胞外配体‑受体信号识别区、跨膜链接区和胞内信号转导区。胞外配体‑受体信号识别区可插入用于细胞识别的蛋白质配体;跨膜链接区连接胞外配体‑受体信号识别区和胞内信号转导区;胞内信号转导区包括转录调控区域和荧光信号报告区域。信号激活质粒包含一个可激活的荧光报告基因。本发明可应用于活细胞表面蛋白原位检测,活细胞间相互作用检测,通过流式细胞分选技术对待测配体进行活性检测及分析等。因此,本发明的工程质粒系统在细胞工程化中具有广阔的应用前景。
Description
技术领域
本发明属于恶性肿瘤的生物治疗技术领域,其涉及到一种用于直接通过细胞-细胞膜表面蛋白分子相互作用进行最佳配体受体组合筛选、优化、和定量检测技术的工程质粒系统及其制备和应用。
背景技术
免疫细胞具有监测和感知身体异常的能力——包括病理性损伤或体内稳态偏离,从而启动保护性和恢复性程序。例如,T细胞可以通过体内运输来感知疾病,并启动有效的反应来消除感染或杀伤肿瘤细胞。原则上,T细胞还具有对疾病产生长期记忆的能力,针对同一抗原可重复被激活。对疾病的监控和反应能力使T细胞成为生物治疗领域中细胞治疗的一个举足轻重的平台。
目前,用于癌症治疗的工程T细胞疗法的主要进展集中于定向天然T细胞对疾病细胞的反应。T细胞可以利用肿瘤特异性T细胞受体 (TCRs)或嵌合抗原受体(CARs)识别新的疾病抗原。目前一系列动物试验和临床试验都证明了CAR-T细胞对血液肿瘤和实体瘤治疗的有效性和安全性,CAR-T细胞在肿瘤的临床治疗中具有巨大的应用潜力和发展前景,是未来治愈肿瘤的利器。
T细胞通过细胞表面的肿瘤特异性T细胞受体(TCRs)或嵌合抗原受体(CARs)识别并杀伤肿瘤细胞,因此,跨膜受体蛋白是介导这类细胞免疫治疗的枢纽。Notch蛋白是一类天然跨膜受体蛋白,其主要功能是介导细胞之间相互信号交流(Natalie L.Stephensonand Johanna M.Avis PNAS October 9,2012 109(41)E2757-E2765)。人工跨膜受体蛋白即以这种基本相互作用为基础将胞外信号识别区和胞内信号转导区人工重组。以Notch蛋白为基础将细胞外传感器模块和细胞内转录模块被异种蛋白结构域取代后形成的新蛋白可以作为产生新的细胞-细胞接触信号通路的通用平台。
肿瘤相关抗原结合区是CAR成功的关键,但是用纯化抗原筛选到的抗体能成功在CAR-T等细胞免疫治疗技术中应用的比例很低,其中原因之一是抗体与抗原的构象受到细胞膜表面结构的影响,细胞表面的相互作用与溶液中的抗体-抗原相互作用方式不同,因此结合能力也不同,因此,基于活细胞-细胞相互作用直接识别和检测抗原并筛选其相应抗体代表了一种更为有效的抗体和嵌合抗原受体筛选和定量检测的方法。
发明内容
本发明的目的是通过搭建一个新的细胞-细胞接相互作用信号转导技术的通用平台,用于高通量配体受体的筛选、优化和定量分析的技术方法及下游应用。这种方法的关键是将单链抗体或单链抗体组合库构建到细胞膜表面的展示载体中,并可通过跨膜区连接一个胞内的信号区进行旁分泌型检测和筛选。
为了达到上述目的,本发明通过设计相关的引物合成目的基因片段,然后将目的基因片段连接到配体-受体信号识别的载体质粒上,将重组质粒导入大肠杆菌工程菌中扩增并提取质粒得到足够拷贝的重组质粒,加入到宿主细胞中,采用慢病毒转染系统或其他细胞转染系统将重组质粒转入到宿主细胞中,完成人工重组质粒构建。
具体包括以下步骤:
·目的基因的合成:基于不同识别域的配体,包含单链抗体、纳米抗体、细胞因子和标签蛋白等受体根据需要定制,设计相应的引物扩增得到对应的目的基因片段。
·重组质粒的构建及验证:将目的基因片段连接到配体-受体信号识别的载体质粒上,将重组质粒导入大肠杆菌工程菌中扩增并提取质粒得到足够拷贝的重组质粒,并通过Sanger测序验证插入序列的正确性。
·细胞转染:采用慢病毒转染或者电转的细胞转染方法将重组质粒转入宿主细胞中。
该工程质粒系统(或工程质粒组)应用于细胞表面配体展示和细胞-细胞相互识别,包含配体-受体信号识别质粒和信号激活质粒,其中配体-受体信号识别质粒包含胞外配体-受体信号识别区、跨膜链接区和胞内信号转导区;所述的胞外配体-受体信号识别区可插入用于细胞识别的蛋白质配体,所述的跨膜链接区用TMC表示,连接胞外配体-受体信号识别区域和胞内信号转导区域;包括core1或core2序列。
core1的碱基序列包括SEQ ID NO:1或该序列内一个或少数几个碱基突变的SEQID NO:1;core1的氨基酸序列包括SEQ ID NO:2或该序列内一个或少数几个氨基酸突变的SEQ ID NO:2。
core2的碱基序列包括SEQ ID NO:3或该序列内一个或少数几个碱基突变的SEQID NO:3;core2的氨基酸序列包括SEQ ID NO:4或该序列内一个或少数几个氨基酸突变的SEQ ID NO:4。
所述的胞内信号转导区包括转录调控区域和荧光信号报告区域,分别用胞内信号转导区和荧光蛋白1表示。
所述的信号激活质粒包含一个荧光信号报告基因,用荧光蛋白2表示;其中荧光蛋白1和2的激发波长不同。
所述的胞外配体-受体信号识别区包括基于不同识别域的配体,所述的不同识别域包含单链抗体、纳米抗体、细胞因子和标签蛋白等构成的配体识别区。
构建好的工程质粒系统应用于基于细胞-细胞相互作用的CD19表面抗原的识别验证,所述的验证实验包含以下步骤:
·细胞转染:将带有CD19抗体的配体-受体信号识别质粒转入宿主细胞中;
·转染实验验证:质粒转染成功的细胞显示荧光蛋白1的相应荧光信号,通过流式分选技术筛选在荧光蛋白1的激发波长下表现相应荧光的细胞,即为包含有人工重组质粒的细胞群;
·细胞-细胞相互作用:将转染成功的细胞群和抗原提呈细胞共同过夜培养,其中系统激活成功的细胞显示荧光蛋白2的相应荧光信号,通过流式分选技术筛选在荧光蛋白2的激发波长下表现相应荧光的细胞;
·CD19抗原识别验证:通过分析表现荧光蛋白2的荧光信号的细胞和表现荧光蛋白1的荧光信号的细胞比例判断CD19抗体对CD19 抗原的识别能力。
附图说明
图1表示本发明中使用的人工重组载体的结构图。
图2表示基于细胞-细胞相互作用的CD19抗原抗体识别验证荧光信号结果。
图3a表示阳性对照FMC63识别CD19的流式分选结果,图3b表示待测样品识别CD19的流式分选结果,图3c表示荧光蛋白2的荧光信号的细胞和表现荧光蛋白1的荧光信号的细胞比例结果。
具体实施方式:
术语定义
受体是指可以识别和选择性地与不同激素、神经递质、药物或细胞内信号分子发生特异性配体-受体结合反应,产生相应地生物效应,引起细胞功能变化的生物大分子。
配体是指对受体具有识别能力并能与之结合的物质。在受体介导的内吞活动中,配体与细胞质膜上受体蛋白结合,激活或启动一系列生物化学反应,最后导致该信号物质的生物效应。细胞膜表面受体一般有胆碱受体、肾上腺素受体和多巴胺受体等。
单链抗体(single-chain variable fragment,scFv)是由抗体重链的可变区与轻链的可变区在一段肽链linker的连接下构成的小分子,是具有抗体活性的最小功能结构单位。scFv能较好地保留其对抗原的亲和活性,并具有分子量小、穿透力强和抗原性弱等特点,在靶向治疗、影像诊断、细胞内免疫、生物检测等方面有着应用。
荧光可以应用于非破坏性跟踪分析生物分子的方法。具体是将蛋白或其他成分被外源性荧光标记,在特定频率激发荧光。
免疫荧光技术(Immunofluorescence technique)——荧光抗体技术。免疫学的基本反应是抗原-抗体反应。由于抗原抗体反应具有高度的特异性,所以当抗原抗体发生反应时,只要知道其中的一个因素,就可以查出另一个因素。免疫荧光技术就是将不影响抗原抗体活性的荧光色素标记在抗体(或抗原)上,与其相应的抗原(或抗体)结合后,在荧光显微镜下呈现一种特异性荧光反应。
FMC63是上个世纪通过动物免疫获得抗CD19的鼠源抗体。 FMC63-scFv已经成功被应用于抗CD19CAR构造,在B细胞急性淋巴细胞白血病的临床试验取得了较好的结果。
CAR嵌合抗原受体是指嵌合抗原受体(chimeric antigen receptor, CAR)是人工设计的可以识别特定蛋白质(抗原)的细胞表面受体。由胞外抗原
结合区、跨膜区域和胞内信号转导区组成。其中跨膜区由来源于单抗克隆抗体的轻链VL和重链VH组成,中间由带韧性的铰链区连接形成单链抗体(single chain fragmentvariable,scfvs)。细胞胞内信号区搭载不同的细胞因子起到免疫细胞激活作用。第一代CARs包含单个胞内激活信号CD3ζ或FcεRIγ。第二代CARs在一代基础上引入了一个共刺激分子,延长了T细胞持续增殖的时间,以分泌更多的细胞因子,从而提高了肿瘤杀伤效力。第三代CARs中则搭载了多个共刺激因子,如 CD28、CD134(OX40)和CD137(4-1BB)等,进一步细化了T细胞中激活的信号通路,延长T细胞增殖活性、存活周期、增强细胞因子分泌等方面的能力;第四代CARs在第三代的基础上增加了可选择性的标记及编码CARs扩增、自杀的启动子。
CAR-T嵌合抗原受体T细胞(Chimeric Antigen Receptor T-Cell),是通过将CAR的DNA序列——识别肿瘤相关抗原的单链抗体和可活化T细胞的胞内信号域在体外进行基因重组,生成重组质粒,再在体外通过转染技术转染处理得到的患者T细胞。CAR-T细胞表达肿瘤抗原受体,经过纯化和大规模扩增后回输到患者体内,具有对特定肿瘤抗原高度亲和性及对抗原负载细胞高效杀伤的特性。
恶性肿瘤是指正常细胞在致癌因素的作用下在局部组织细胞增生所形成的异常新生物,而其在结构功能和代谢方面良性肿瘤形成明显的区别。相较于良性肿瘤,恶性肿瘤与原组织形态差异大、分化水平低、能无限增殖、拥有更快的生长速度、浸润性和转移能力。
免疫细胞(immune cell),是指参与免疫应答或与免疫应答相关的细胞。包括先天性淋巴细胞淋巴细胞、树突状细胞、单核/巨噬细胞、粒细胞、肥大细胞和能识别抗原、产生特异性免疫应答的淋巴细胞等,以及它们的前体细胞等,是免疫系统的功能单元。
TCRs,T细胞受体(T cell receptor,TCR)是T细胞表面的特异性受体,负责识别由主要组织相容性复合体(MHC)所呈递的抗原,与 B细胞受体不同,并不能识别游离的抗原。通常情况下,T细胞受体与抗原间拥有较低的亲和力,因而同一抗原可能被不同的T细胞受体所识别,而同一受体可能识别许多种抗原。
部分材料来源说明于此
SfiI酶(NEB公司,R0123L)
CIP(NEB公司,M0290S)
去内毒质粒小提试剂盒(Omega公司,D6950-02)
HEK293(ATCC)
DMEM(Life,11966025)
FBS(Gibco,10099141)
polybrene(Merck,TR-1003-G)
流式细胞仪(Beckman,Cytoflex S)
实验案例一重组质粒的构建
通过设计引物合成目的基因片段,然后将目的基因片段连接到配体 -受体信号识别的载体质粒上,将重组质粒导入大肠杆菌工程菌中扩增并提取质粒得到足够拷贝的重组质粒,通过Sanger测序验证插入序列的正确性。将验证正确的重组质粒加入到HEK293细胞中,采用慢病毒转染系统或其他细胞转染系统将重组质粒转入到HEK293细胞中,完成人工重组质粒构建。
具体步骤如下:
1构建CD19antibody信号识别质粒:
·酶切:取2μg配体-受体信号识别质粒,加入1μlSfiI酶(NEB公司,R0123L)进行酶切,50℃酶切2h后加入1μl CIP (NEB公司,M0290S)去磷酸化处理20min,PCR纯化回收酶切产物;
·酶切:取2μg CD19的抗体FMC-63质粒,加入1μl SfiI进行酶切,50℃酶切2h后胶回收酶切产物;
·连接:配体-受体信号识别质粒载体sfiI酶切产物和FMC-63 sfiI酶切产物各取50ng,16℃连接1h;
·转化:取2μl连接产物转入100μl大肠杆菌DH5α感受态细胞中,冰上孵育30min,42℃热激45s,冰上再放5min,加入2ml的 LB培养基,37℃培养1h;
·涂板:离心收集菌体,加入100μl的培养基重悬,全部涂布到含有氨
苄青霉素的LB平板中,37℃培养过夜;
·挑克隆:随机挑选3个单克隆到含有3ml LB培养基及100 ng/μl氨苄青霉素的14ml培养管中,37℃过夜培养;
·质粒提取:离心收集过夜生长的菌体,用去内毒质粒小提试剂盒(Omega公司,D6950-02)进行质粒提取,Sanger测序法验证序列的正确性。
实验结果:Sanger测序结果显示目的基因序列成功插入配体-受体信号识别的载体质粒中,如图1所示,成功获得具有图中核心组件的人工重组质粒。
实验案例二包含重组质粒的活细胞荧光验证
通过将带有特定配体的信号识别质粒转入HEK293宿主细胞中;质粒转化成功的细胞显示红色荧光,通过流式分选技术筛选在555nm的激发光下表现红色荧光的细胞,即为包含有配体-受体信号识别质粒的细胞群;
具体步骤如下:
·慢病毒包装
准备包装CD19antibody信号识别质粒和信号激活质粒病毒。
1.铺一块6孔板,每孔加5x105个293T细胞,混匀在1ml DMEM (Life,11966025)+10%FBS(Gibco,10099141)中;
2. 24h后,准备包装病毒,取1.5μg CD19antibody信号识别或信号激活质粒,0.6μg PMD2G,0.9μg PSPAX,6μl P3000和125μl DMEM加到A管中;
3.取5μl lipo3000(Life,L3000015)和125μl DMEM加到B管中,室温放置5min;
4.B管溶液全部加到A管,混匀,室温放置5-15min;
5.将混合溶液加到6孔板1个孔中,;
6. 6h后,吸掉样品孔培养基,加入3ml DMEM+10%FBS;
7. 72h后,收集包装的病毒。使用Lenti-X p24Rapid Titer Kit (clontech,632200)试剂盒测定病毒滴度。
·慢病毒转染
用慢病毒转染的方法将CD19antibody信号识别质粒和信号激活质粒表达在HEK293细胞上。
1.HEK293细胞计数,计数后取5x105个HEK293细胞,1000rpm 离心5min,用1mlDMEM+10%FBS的培养基重悬;
2.对样品管加MOI=2的CD19antibody-TM和信号激活病毒,补齐 DMEM+10%FBS培养基至1ml;
3.样品管加polybrene(Merck,TR-1003-G)至终浓度为8μg/ml,混匀,对应加到12孔板;
4. 12孔板放在离心机800g离心1h,细胞培养箱中过夜培养;
5. 24h后将样品孔细胞转移到10cm细胞培养皿继续培养,培养过程中,使用荧光显微镜观察细胞荧光表达情况;
实验结果:导入工程质粒的细胞在荧光显微镜下表现出红色荧光,说明重组质粒成功导入到宿主细胞中。
实验案例三CD19表面抗原的识别
细胞膜表面CD19抗原与FMC-63、待测样品识别能力的对比分析
将CD19过表达的Raji细胞分别与表达CD19antibody(FMC-63、待测样品)的报告蛋白的HEK293细胞共孵育,然后使用流式细胞仪分析RFP和BFP表达。
1.将表达荧光报告蛋白的HEK293细胞消化,1000rpm离心 5min,用DMEM+10%FBS重悬至1.5x104个HEK293细胞/ml,加1ml 重悬后的HEK293细胞到24孔板一个孔中,37℃,5%CO2放在细胞培养箱中过夜培养24h;
2.Raji细胞样品1000rpm离心5min,用DMEM+10%FBS重悬至 1x105个细胞/ml,加入HEK293的细胞中,共同抚育培养;
3. 24-72h后消化细胞,用流式缓冲液洗涤2次,0.5ml流式缓冲液重悬后使用流式细胞仪分析(Beckman,Cytoflex S)。
实验结果:如图2、3a、3b、3c所示,质粒转染成功的宿主细胞表现为红色荧光信号,而能够通过细胞-细胞相互作用被CD19抗原识别的宿主细胞表现蓝色荧光信号。比对表现蓝色荧光信号的细胞和表现红色荧光信号的数量比例说明阳性对照的FMC63和CD19的结合能力强于待测样品。本发明的人工重组质粒系统能够实现细胞层面的配体-受体识别和结合,具有很好的应用前景。
综上所述,本发明的应用于细胞表面配体展示和细胞-细胞相互识别的人工重组质粒系统,可以用于细胞-细胞膜表面展示的配体-受体组合筛选、优化、和定量的检测技术。同时,该系统本发明可应用于活细胞表面蛋白原位检测,活细胞间相互作用检测,通过流式细胞分选技术对待测配体进行活性检测及分析等。因此,本发明的人工重组质粒系统在细胞工程化中具有广阔的应用前景。
尽管本发明的内容已经通过上述优选实施例作了详细介绍,但应当认识到上述的描述不应被认为是对本发明的限制。在本领域技术人员阅读了上述内容后,对于本发明的多种修改和替代都将是显而易见的。因此,本发明的保护范围应由所附的权利要求来限定。
序列表
<110> 常州费洛斯药业科技有限公司
<120> 用于活细胞间膜蛋白展示和相互作用检测的工程质粒系统
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ccctggcaga actgcagcgc ggccttgcaa tgttggcgat acttcaatga cggaaagtgt 240
gatgaacaat gtgcaacggc tggttgtctc tatgatgggt ttgactgcca aagattggaa 300
gggcaatgca atcccctcta cgatcagtat tgccgcgatc actatgctga tggacactgc 360
gatcaggggt gcaacaacgc agagtgcgag tgggacggtt tggactgtgc agacgatgtg 420
ccgcaaaaac ttgctgtggg ctcccttgtt ttggtcgttc acataccacc cgacgaactt 480
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tttaggaggg acgcaaacgg agaagcattg atttttccat attacggttc cgaacatgag 600
ctcagcaagc ataaaaggag tgactggacc gaccctggcc aacttatgca aagagctagg 660
cgcagcctca cttccttctt gaaaccccgc actcggcgag aactcgatca catggaagtc 720
aaaggtagca ttgtgtacct cgaaatagat aacagacagt gttttcagca atctgatgag 780
tgttttcaga gtgcaacgga tgtcgctgcc ttcctggggg ccttggcaag tagtggaaac 840
cttaatgtgc cttactgcat cgaggcggta acctgcgaag gcggtccacc aaagacggga 900
gaaatgtatc caatgttctt ggtgcttctg gctcttgccg tcttggcgct tgccgcagtc 960
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Pro Tyr Tyr Gly Ser Glu His Glu Leu Ser Lys His Lys Arg Ser Asp
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Ser Phe Leu Lys Pro Arg Thr Arg Arg Glu Leu Asp His Met Glu Val
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Gln Ser Asp Glu Cys Phe Gln Ser Ala Thr Asp Val Ala Ala Phe Leu
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Gly Ala Leu Ala Ser Ser Gly Asn Leu Asn Val Pro Tyr Cys Ile Glu
275 280 285
Ala Val Thr Cys Glu Gly Gly Pro Pro Lys Thr Gly Glu Met Tyr Pro
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gctccgccct ttagtggcag tcgctgcgag ttgtacacag caccccctag cacaccgccc 60
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gctgaaggta cgctggtcat cgtcgtgttg atgccaccgg agcaactctt gcaggatgca 480
cgctcctttt tgagggctct tggaacgctt ttgcatacta acctccgcat caagcgcgat 540
tcccaaggag agctcatggt atatccatat tatggggaaa agtcagccgc gatgaagaaa 600
cagcggatga ccagacggag cttgccagga gagcaagaac aagaggtagc ggggtctaaa 660
gtcttccttg aaatcgacaa caggcaatgc gtacaagata gtgaccattg cttcaagaat 720
actgacgcgg ctgcggctct tttggcgtca cacgccatac aaggaacact tagttacccc 780
ctcgttagtg tcgtcagcga atcattgaca ccggaacgga ctcagctgct gtgctacctg 840
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aaacgcaaga ggaagcacgg ttcc 924
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<212> PRT
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Asp Asn His Cys Asp Gln Gly Cys Asn Ser Glu Glu Cys Gly Trp Asp
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Ile Lys Arg Asp Ser Gln Gly Glu Leu Met Val Tyr Pro Tyr Tyr Gly
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Pro Gly Glu Gln Glu Gln Glu Val Ala Gly Ser Lys Val Phe Leu Glu
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Ile Asp Asn Arg Gln Cys Val Gln Asp Ser Asp His Cys Phe Lys Asn
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Thr Asp Ala Ala Ala Ala Leu Leu Ala Ser His Ala Ile Gln Gly Thr
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Lys His Gly Ser
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Claims (5)
1.一种用于活细胞间膜蛋白展示和相互作用检测的工程质粒系统,该工程质粒系统包含配体-受体信号识别质粒和信号激活质粒,其中配体-受体信号识别质粒包含胞外配体-受体信号识别区、跨膜链接区和胞内信号转导区;所述的胞外配体-受体信号识别区可插入用于细胞识别的蛋白质配体,所述的跨膜链接区用TMC表示,连接胞外配体-受体信号识别区和胞内信号转导区,包括core1或core2序列;
core1的碱基序列包括SEQ ID NO:1或该序列内一个或少数几个碱基突变的SEQ IDNO:1;core1的氨基酸序列包括SEQ ID NO:2或该序列内一个或少数几个氨基酸突变的SEQID NO:2;
core2的碱基序列包括SEQ ID NO:3或该序列内一个或少数几个碱基突变的SEQ IDNO:3;core2的氨基酸序列包括SEQ ID NO:4或该序列内一个或少数几个氨基酸突变的SEQID NO:4;
所述的胞内信号转导区包括转录调控区域和荧光信号报告区域,分别用胞内信号转导区和荧光蛋白1表示;
所述的信号激活质粒包含一个荧光信号报告基因,用荧光蛋白2表示;其中荧光蛋白1和2的激发波长不同。
2.根据权利要求1所述的工程质粒系统,所述的胞外配体-受体信号识别区基于不同识别域的配体,所述的不同识别域包含单链抗体、纳米抗体、细胞因子和标签蛋白构成的配体识别区。
3.根据权利要求1所述的工程质粒系统,通过以下方法制得:通过设计引物合成目的基因片段,然后将目的基因片段连接到配体-受体信号识别的载体质粒上,将构建的重组质粒导入大肠杆菌工程菌中扩增并提取质粒,得到足够拷贝数的重组质粒,然后采用慢病毒转染系统或其他细胞转染系统将重组质粒转入到宿主细胞中,完成人工重组质粒构建。
4.根据权利要求3所述的工程质粒系统,所述的方法包括以下步骤:
目的基因的合成:基于不同识别域的配体,包含单链抗体、纳米抗体、细胞因子和标签蛋白受体根据需要定制,设计相应的引物扩增得到对应的目的基因片段;
重组质粒的构建及验证:将目的基因片段连接到配体-受体信号识别的载体上,将重组质粒导入大肠杆菌工程菌中扩增并提取质粒得到足够拷贝的重组质粒,并通过Sanger测序验证插入序列的正确性;
细胞转染:采用慢病毒转染或者电转的细胞转染方法将重组质粒转入宿主细胞中。
5.根据权利要求1所述的工程质粒系统的应用,包括基于细胞-细胞相互作用的CD19表面抗原的识别验证,所述的验证实验包含以下步骤:
细胞转染:将带有CD19抗体的信号识别质粒转入宿主细胞中;
转染实验验证:质粒转染成功的细胞显示荧光蛋白1的相应荧光信号,通过流式分选技术筛选在荧光蛋白1的激发波长下表现相应荧光的细胞,即为包含有人工重组质粒的细胞群;
细胞-细胞相互作用:将转染成功的细胞群和抗原提呈细胞共同过夜培养,其中系统激活成功的细胞显示荧光蛋白2的相应荧光信号,通过流式分选技术筛选在荧光蛋白2的激发波长下表现相应荧光的细胞;
CD19抗原识别验证:通过分析表现荧光蛋白2的荧光信号的细胞和表现荧光蛋白1的荧光信号的细胞比例判断细胞膜表面展示的CD19抗体对CD19抗原的识别能力。
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Cited By (2)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN111269941A (zh) * | 2020-02-27 | 2020-06-12 | 南京鼓楼医院 | 一种基于双色荧光系统的活化car-t细胞的示踪及定量方法 |
CN113881701A (zh) * | 2021-09-08 | 2022-01-04 | 山东大学 | 一种阳性转染细胞亲和分选方法及试剂盒 |
Citations (2)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN108064283A (zh) * | 2015-02-24 | 2018-05-22 | 加利福尼亚大学董事会 | 结合触发的转录开关及其使用方法 |
TW201905198A (zh) * | 2017-06-19 | 2019-02-01 | 美商細胞設計實驗室股份有限公司 | 用於降低嵌合刻痕受體之免疫原性的方法與組合物 |
-
2019
- 2019-04-16 CN CN201910305032.5A patent/CN111826398A/zh active Pending
Patent Citations (2)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN108064283A (zh) * | 2015-02-24 | 2018-05-22 | 加利福尼亚大学董事会 | 结合触发的转录开关及其使用方法 |
TW201905198A (zh) * | 2017-06-19 | 2019-02-01 | 美商細胞設計實驗室股份有限公司 | 用於降低嵌合刻痕受體之免疫原性的方法與組合物 |
Non-Patent Citations (3)
Title |
---|
NCBI: "NCBI: PNI26471.1" * |
NCBI: "NCBI:NP_571516.1" * |
ROYBAL ET AL.: "Engineering T Cells with Customized Therapeutic Response Programs Using Synthetic Notch Receptors" * |
Cited By (3)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN111269941A (zh) * | 2020-02-27 | 2020-06-12 | 南京鼓楼医院 | 一种基于双色荧光系统的活化car-t细胞的示踪及定量方法 |
CN113881701A (zh) * | 2021-09-08 | 2022-01-04 | 山东大学 | 一种阳性转染细胞亲和分选方法及试剂盒 |
CN113881701B (zh) * | 2021-09-08 | 2024-04-19 | 山东大学 | 一种阳性转染细胞亲和分选方法及试剂盒 |
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