CN117106788A - 一种文昌鸡羽色调控相关基因及其应用 - Google Patents
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Abstract
本发明公开了一种文昌鸡羽色调控相关基因及其应用，所述基因的核苷酸序列如SEQ ID NO.1所示，研究表明该基因与文昌鸡羽色紧密相关。本发明还具体提供了一个与羽色相关的SNP位点，所述基因和SNP位点可用于文昌鸡分子标记辅助育种，为文昌鸡羽色分子选育奠定基础。
Description
技术领域
本发明涉及文昌鸡育种技术领域，具体涉及一种文昌鸡羽色调控相关基因及其应用。
背景技术
家鸡(Gallus gallus domesticus)是世界上养殖范围最广、数量最多的家养动物。我国幅员辽阔、地形地貌多样、气候多变，形成了不同的地域民族文化，孕育了具有不同特性的地方鸡品种。文昌鸡是我国热带地区的优良地方鸡种，因发源于海南省文昌市而得名。其形成历史悠久，据传在清代，家鸡由福建、广东地区的移民带入海南岛，在文昌市潭牛镇天赐村，家鸡啄食了榕树籽，变得体质极佳，深受村民喜爱。在当地特定的热带季风海洋性气候条件下，经长期选育，逐渐形成了有别于其他家鸡品种的身材娇小、肉质鲜嫩、肉香浓郁、皮薄骨酥的优质文昌鸡。
对羽色进行选育是现代家鸡养殖业的一个重要课题。同其他鸟类一样，羽毛是家鸡皮肤最复杂的附属物之一，其颜色、结构和分布复杂多样，能够为家鸡提供保持体温、信息交流、外形伪装等重要价值。家鸡羽色也具有经济价值，例如，白羽相比于黑羽、红羽等其他颜色，屠宰后绒毛和毛根残留几乎不会引起注意，鸡肉卖相更好，所以白羽肉鸡是肉鸡工业生产的首选。
羽色是目前文昌鸡选育的一个重要标志。不同的文昌鸡羽色可能由不同基因控制。通过对羽色相关调控基因的分析，可以在育种早期筛选出目标羽色的个体。但由于羽色调控机制复杂，目前关于文昌鸡调控相关基因和特异性变异位点的研究并不多见。
发明内容
鉴于现有技术的不足，本发明提供了一种文昌鸡羽色调控相关基因及其应用。
本发明选取经过近20年选育的9个文昌鸡群体为研究对象，进行全基因组重测序，解析文昌鸡的群体遗传结构和遗传多样性，通过全基因组关联分析和全基因组选择信号分析，鉴定文昌鸡羽毛颜色的相关位点和基因。
本发明涉及如下几个方面：
本发明的目的之一是提供一种文昌鸡羽色调控相关基因，所述基因的核苷酸序列如SEQ ID NO.1所示。所述基因编码的蛋白的氨基酸序列如SEQ ID NO.2所示。
另一方面，本发明还提供所述基因或所述蛋白在文昌鸡分子辅助育种中的应用，具体为在羽色选育中的应用。
具体的：所述应用包括：SEQ ID NO.1所示序列的第64位存在一个SNP位点，第64位的S为碱基C或G；检测所述SNP位点信息，当待测文昌鸡的基因组中该SNP位点的基因型为CC型时，则待测样本的羽色为白色的概率高于其他羽色。
进一步的，上述SNP分子标记用于文昌鸡白羽品种育种的步骤为：取候选亲本的DNA，以提取的DNA为模板进行PCR扩增，产物测序，筛选出前述SNP位点基因型为CC纯合的亲本进行育种。
与现有技术相比，本发明的有益效果是：
本发明提供了一个文昌鸡羽色调控相关基因和SNP分子标记，可用于文昌鸡分子标记辅助育种，为文昌鸡羽色分子选育奠定基础。
附图说明
图1：文昌鸡白羽-黄羽全基因组关联分析结果的曼哈顿图。虚线为Bonferroni校正阈值线，虚线上方位点为显著位点。
具体实施方式
下面通过具体实施例对本发明作进一步的描述，以使本领域的普通技术人员能够充分的理解本发明的技术内容。
实施例1
1.1实验群体重测序
选择不同羽色的文昌鸡群体，每个群体随机选择三代内无直接亲缘关系的雌雄样本各10个。所有群体选育和样本采集由海南省农业科学院畜牧兽医研究所顾丽红老师团队完成。从鸡血卡样品中提取基因组DNA，基于华大MGI-2000/MGI-T7测序平台，构建小片段文库(350bp)进行双末端测序，获得全基因组重测序数据。所有实验均按《实验动物管理规定》(中国科学技术部，2017年3月修订)执行，并经海南省农业科学院实验动物中心机构动物保护与使用专业委员会批准(批准编号：HNXMSY-20210533)。
1.2变异位点检测与质控
为了保证数据分析质量，使用软件Fastp(version 0.20.0)对raw reads进行过滤，数据处理的步骤如下：去除接头序列(adapter)；当测序read中含有的N的含量超过该条read长度比例的10％时，去除此对paired reads；当测序read中含有的低质量(Q≤20)碱基数超过该条read长度比例的40％时，去除此对paired reads。
1.3变异检测
使用软件BWA-MEM(version 0.7.17)将质控后的clean reads与参考基因组GRCg7b(https://www.ncbi.nlm.nih.gov/assembly/GCF_016699485.2/)进行比对。根据clean reads在参考基因组的比对结果，使用软件GATK(version 4.2.1.0)的HaplotypeCaller模块进行变异检测。
1.4变异位点质控
使用软件GATK(version 4.2.1.0)的VariantFiltration模块进行过滤。
1.5基因组变异位点注释
使用ANNOVAR(version 20191024)软件对所有变异位点进行注释。并根据变异位点在基因组的不同区域以及变异产生的影响分别进行统计。
1.6数据集准备
完成上述数据质控，检测出SNPs和INDELs变异位点，用于变异位点注释。去除所有的INDELs以及性染色体上的SNPs，并只保留双等位基因位点，用于后续文昌鸡羽色的全基因组关联分析和选择信号分析。
1.7采用全基因组关联分析和全基因组选择信号分析鉴定文昌鸡不同羽色的候选基因
对数据集进行关于文昌鸡羽色的分析。进行两两群体间的Case-Control全基因组关联分析；以及一种羽色为Case、其他羽色合并为Control的关联分析。
通过绘制QQ图来判定GWAS结果的可靠性。通过基因组控制(Genomic control,GC)计算膨胀因子(Inflation factor,λ)来检验群体分层。使用Bonferroni校正划定阈值，确定显著关联位点。对确定出显著位点较少的关联分析采用错误发现率法(FDR)再次确定显著位点，阈值为0.05。
在白羽-黄羽和白羽-其他全基因组关联分析中，达到阈值的位点注释到SEQ IDNO.1所示的基因，该基因与文昌鸡的白羽性状相关。基因第64位为错义突变(G>C)，是影响基因的重要候选位点，该位点定位在基因的1号外显子，是三联密码子的第一个碱基，导致氨基酸从天冬氨酸转变为组氨酸。
实施例2
对实施例1筛选出的位点，以不同羽色的文昌鸡DNA为模板，利用PCR技术进行扩增，扩增产物测序，鉴定该位点的基因型，统计基因型频率。结果见下表。
表1
该位点在白羽群体的基因型频率为GG(13.2％)、GC(16.8％)和CC(70％)，在黄羽群体的基因型频率为GG(32.6％)、GC(43.2％)和CC(24.2％)。结果表明该位点与白羽性状显著关联，该位点可用于预测文昌鸡羽色。当待测文昌鸡的基因组中该位点的基因型为CC型，则待测样本的羽色为白色的概率高于其他羽色。可见本发明提供的基因及候选位点可作为文昌鸡分子辅助育种的工具，用于文昌鸡羽色的选育。
实施例3
实施例1所述SNP分子标记可用于文昌鸡品种选育，具体为白羽品种的选育。文昌鸡白羽品种育种的步骤为：取候选亲本的DNA，以提取的DNA为模板进行PCR扩增，产物测序，筛选出SNP位点基因型为CC纯合的亲本进行育种，则其后代为白羽品种的概率明显提高。
以上仅为本发明的部分实施案例，并不用以限制本发明本发明的范围，凡在本发明的精神和原则之内，对本发明方案所做的修改、等同替换等，均属于本发明的保护范围之内。
Claims (6)
1.一种文昌鸡羽色调控相关基因，其特征在于，所述基因的核苷酸序列如SEQ ID NO.1所示。
2.权利要求1所述基因编码的蛋白，其特征在于，所述蛋白的氨基酸序列如SEQ IDNO.2所示。
3.权利要求1所述基因或权利要求2所述蛋白在文昌鸡分子标记辅助育种中的应用。
4.权利要求1所述基因或权利要求2所述蛋白在文昌鸡羽色选育中的应用。
5.根据权利要求4所述的应用，其特征在于，SEQ ID NO.1所示序列的第64位存在一个SNP位点，第64位的S为碱基C或G；检测所述SNP位点信息，根据SNP位点的基因型预测文昌鸡羽色。
6.根据权利要求5所述的应用，其特征在于，当待测文昌鸡的基因组中该SNP位点的基因型为CC型时，则待测样本的羽色为白色的概率高于其他羽色。
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