| 减数分裂关键基因Spo11和Mlh1在鲫鲤杂交鱼中的遗传分析 |
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| 引用本文： | 朱辣,张美文,朱明,李琪,贺旺超,秦伟玲,周毅,杨聪慧,张纯,刘少军.减数分裂关键基因Spo11和Mlh1在鲫鲤杂交鱼中的遗传分析[J].南方农业学报,2021,52(8):2251-2258. |
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| 作者姓名： | 朱辣 张美文 朱明 李琪 贺旺超 秦伟玲 周毅 杨聪慧 张纯 刘少军 |
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| 作者单位： | 省部共建淡水鱼类发育生物学国家重点实验室/多倍体鱼繁殖与育种技术教育部工程研究中心/湖南师范大学生命科学学院, 长沙 410081 |
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| 基金项目： | 国家自然科学基金项目（31873038，31730098，U19A2040）；国家现代农业产业技术体系建设专项（CARS-45） |
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| 摘 要： | 【目的】明确鲫鲤杂交鱼减数分裂关键基因（Spo11和Mlh1）的序列和表达特征，揭示异源双亲减数分裂因子在杂交品系的传代中是否存在选择压力，为解析远缘杂交跨越生殖障碍打下遗传学基础。【方法】以已形成稳定品系的鲫鲤杂交鱼为研究对象，包括红鲫（RCC）、湘江野鲤（CC）、鲫鲤杂交子代（F1）和异源四倍体鲫鲤（4nAT），通过分子克隆、序列比对及Western blotting等方法研究Spo11基因和Mlh1基因在鲫鲤杂交鱼中的遗传规律和表达特征，并分析其与原始父母本间的关系。【结果】RCC、CC、F1和4nAT的Spo11基因CDS序列绝大部分为1152 bp，仅4nAT中存在1157 bp的CDS序列；在F1和4nAT中均稳定获得2种原始亲本的独立遗传片段，且在4nAT中还检测到原始父母本重组基因片段［4n-SPO11-11-2（MT648223）］，其与原始母本RCC-Spo11基因对应片段的相似性为96.2%，与原始父本CCSpo11基因对应片段的相似性为93.7%，表明Spo11基因表达模式并非完全保守性及4nAT存在遗传变异性。Mlh1基因CDS序列在鲫鲤杂交鱼中同样稳定存在，表现出稳定的杂合遗传特征，即F1和4nAT中既存在RCC-Mlh1基因CDS序列（相似性为99.5%），也存在CC-Mlh1基因CDS序列（相似性为99.4%），鲫鲤杂交鱼相应的遗传片段和原始亲本间仅存在极少的基因位点变异。Western blotting检测结果表明，SPO11蛋白和MLH1蛋白在鲫鲤杂交子代中均能正常表达。其中，4nAT的精巢中仅表达相对分子量较大的SPO11-β亚体，而F1的精巢中仅表达相对分子量较小的SPO11-α亚体。【结论】F1和4nAT能稳定杂合遗传原始亲本RCC的Spo11基因和Mlh1基因，虽然在CDS序列结构上存在少量差异，但最终都能独立表达相关蛋白，即杂交鱼形成过程中减数分裂关键基因的表达正常可为其跨越生殖障碍打下分子基础。
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| 关 键 词： | 异源四倍体鲫鲤(4nAT) Spo11基因 Mlh1基因 减数分裂 远缘杂交 遗传模式 |
| 收稿时间： | 2021-06-28 |
Genetic analysis of key genes Spo11 and Mlh1 controlling meiosis in hybrid fish of Carassius auratus red var.(♀)×Cyprinus carpio L.(♂) |
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| ZHU La,ZHANG Mei-wen,ZHU Ming,LI Qi,HE Wang-chao,QIN Wei-ling,ZHOU Yi,YANG Cong-hui,ZHANG Chun,LIU Shao-jun.Genetic analysis of key genes Spo11 and Mlh1 controlling meiosis in hybrid fish of Carassius auratus red var.(♀)×Cyprinus carpio L.(♂)[J].Journal of Southern Agriculture,2021,52(8):2251-2258. |
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| Authors: | ZHU La ZHANG Mei-wen ZHU Ming LI Qi HE Wang-chao QIN Wei-ling ZHOU Yi YANG Cong-hui ZHANG Chun LIU Shao-jun |
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| Affiliation: | State Key Laboratory of Developmental Biology of Freshwater Fish/Engineering Research Center of Polyploid Fish Reproduction and Breeding of Ministry of Education/College of Life Sciences, Hunan Normal University, Changsha 410081, China |
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| Abstract: | 【Objective】 The present study analyzed the characteristics of gene sequences and expression of key genes (Spo11 and Mlh1) in meiosis from the hybrid crucian carp(Carassius auratus red var., ♀, abbreviated as RCC)×common carp(Cyprinus carpio L., ♂, abbreviated as CC), which would reveal whether there was selection pressure for meiosis factors in hybrid lines, providing the genetic basis for understanding the mechanism of breaking through reproductive barriers in hybrid fishes.【Method】 In this study, the hybrid strains of RCC×CC which has formed stable strain was taken as research objects, containing RCC, CC, allodiploid hybrid of RCC(♀)×CC(♂) (abbreviated as F1) and allotetraploid hybrid of RCC(♀)×CC(♂) (abbreviated as 4nAT), respectively. The genetic and expression characteristics of Spo11 gene and Mlh1 gene in hybrid crucian carp were studied by molecular cloning, sequence alignment and Western blot, and the gene-tic relationships among them and their original parents were analyzed.【Result】 Most of the CDS sequences of Spo11 gene in RCC, CC, F1 and 4nAT were 1152 bp, butsome1157bp CDS sequences were also found in 4nAT. Two independent genetic fragments of the original parents were stably obtained in F1 and 4nAT, and the recombinant gene fragment ［4N-SPO11-11-2(MT648223) ］of the original parents was also detected in 4nAT, and its similarity to the corresponding fragment of the RCC-SPO11 gene of the maternal was 96.2%, and to the corresponding fragment of the CC-Spo11 gene of the paternal was 93.7%, indicating that the expression pattern of Spo11 gene was not completely conserved and 4nAT possessed some genetic variations. CDS sequences of Mlh1 gene also were stably in hybrid crucian carp, showing stable heterozygous genetic characteristics, namely both CDS sequences of RCC-MLH1 gene(99.5%) and CC-MLH1 gene (99.4%) in F1 and 4nAT, and there were only a few gene loci variation in the corresponding genetic fragment between the original parents and hybrid fish. Furthermore, the results of western blotting showed that both SPO11protein and MLH1 protein expressed normally in the hybrid progeny of RCC(♀)×CC(♂). The SPO11-β subunit with large molecular weight was only expressed in the testis of 4nAT, while SPO11-α subunit with small molecular weight was only expressed in testis of F1.【Conclusion】 These results show that F1 and 4nAT stabilize the Spo11 gene and Mlh1 gene of the heterozygous inherited original parental RCC, although there are small differences in CDS sequence structure, they can express related protein independently. It indicates that key meiotic genes are expressed normally during hybrid fish formation, and lay an important genetic basis for the analysis of the distant hybridization to overcome reproductive barriers. |
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