【作者】 马冬梅； 朱择敏； 白俊杰； 全迎春； 樊佳佳； 田园园； 廖国礼； 梁旭方；

【Author】 Ma Dong-Mei;Zhu Ze-Mmin;Bai Jun-Jie;Quan Ying-Chun;Fan Jia-Jia;Tian Yuan-Yuan;Liao Guo-Li;Liang Xu-Fang;Key Lab of Tropical & Subtropical Fish Breeding & Cultivation of Chinese Academy of Fishery Sciences, Pearl River Fisheries Research Institute of Chinese Academy of Fishery Sciences;Department of Bioengineering,College of Life Science and Technology,Jinan University;

【机构】 中国水产科学研究院珠江水产研究所农业部热带亚热带水产资源利用与养殖重点实验室； 暨南大学生命科学技术学院生物工程学系；

【摘要】 大口黑鲈(Micropterus salmoides)为肉食性经济鱼类,喜食活饵或冰鲜鱼,养殖过程中大量投喂冰鲜鱼或大量使用鱼粉作为饲料,增加了养殖成本,且容易对环境造成污染。为了降低养殖成本、保护环境,使其适合喂食蛋白含量适中的人工配合饲料,提高大口黑鲈对人工配合饲料的适应能力,选育适合食人工配合饲料的大口黑鲈是解决这个问题的有效途径。有研究表明,饲料中的脂肪水平对大口黑鲈的脂蛋白脂肪酶(lipoprotein lipase,LPL)的表达水平有显著的影响。本研究以大口黑鲈LPL基因作为候选基因,根据已知的c DNA序列设计引物,PCR扩增测序得到大口黑鲈LPL type1基因组序列6 170 bp,包括10个外显子和9个内含子,LPL type2基因组序列4 419 bp,包括5个外显子和4个内含子。利用直接测序法,在LPL type1基因中筛选到11个SNP位点,在LPL type2中筛选到6个SNP位点。将一个喂食冰鲜鱼的成鱼养殖群体中的192尾鱼和一个幼鱼驯食养殖群体中的142尾鱼,分别用筛选得到的SNP位点进行基因分型,结果发现在LPL type1基因中筛选到的11个SNP位点紧密连锁,用T+156A表示;在LPL type2基因中筛选到的6个SNP位点也紧密连锁,用C-224T表示,而LPL type1和LPL type1的SNP位点间不存在连锁关系,且SNP位点所构成的基因型在两个群体中的分布频率相似。将上述SNP位点所构成的基因型与两个大口黑鲈群体的体重、全长和体高等生长性状进行相关性分析,结果表明,LPL基因中的SNP位点虽然与成鱼的生长性状不存在显著的相关性,但在LPL type1基因T+156A SNP位点的AA基因型与TT基因型与幼鱼在驯食后的饱食与空腹状态存在显著的相关性,且AA基因型在幼鱼体质量和全长两个性状上存在显著的优势(P<0.05),说明T+156A SNP位点与大口黑鲈适应人工配合饲料的能力有显著相关。LPL type1基因可作为提高大口黑鲈成活率、选育适合食人工配合饲料大口黑鲈的潜在候选基因。本研究为推进大口黑鲈适应人工配合饲料新品种的选育工作提供了可靠的研究数据,也为其它肉食性鱼类的驯食选育工作提供了借签和参考。更多还原

【Abstract】 Largemouth bass(Micropterus salmoides) is a very important commercial farming fish with the annual production of more than 340,000 tons in China. But it is a carnivorous teleost, and a large amount of forage fish and fish meal were used as food for largemouth bass very year. The residual forage fish and fish meal not only increase the culture costs, but also pollute waterbodies and farmlands seriously. In order to reduce culture costs and protect the natural environment, the selective breeding of the largemouth bass strain suitable for formulated feeds was carried out. It is a way to improve the effective conversion capability of largemouth bass for formulated feeds. In the previous study, the lipid levels of the formulated feeds have significant effects on the expression levels of two lipoprotein lipase genes(LPL type1 and LPL type2) in livers of largemouth bass. The report suggests the two genes potentially related to the adaptability of largemouth to formulated feed. In the current study, the primers LPL genes were designed according the known c DNA and the PCR amplifications were performed with largemouth bass genome DNAs as templates. After amplifying and sequencing, the genomic sequences of LPL type1 gene with the length of 6170 bp were identified including 10 exons and 9 introns, and LPL type2 gene with the length of 4419 bp were identified including 5 exons and 4 introns. By use of direct sequencing, eleven SNPs were detected in LPL type1 gene, and six SNPs were detected in LPL type2 gene. A total of 192 individuals from the cultured adult fish population fed on iced fresh fish and a total of 142 individuals from the cultured juvenile fish population fed on formulated feed were genotyped according to the SNP sites. The results showed that the eleven SNPs in LPL type1 closely linked, and were named as T+156A. As well as the six SNPs in LPL type2 were closely linked, and were named as C-224 T. But the SNP sites were not closed linked between the two genes. The genotype frequency and gene frequency of the SNPs were similar between the two populations. And the calculations showed that the two populations were in Hardy-Weinberg equilibrium for the two SNPs. The general linear model was applied to analyze the relationship between SNPs and the full/empty traits, as well as the growth traits of largemouth bass. The results showed that there were no significant association between the LPL SNP sites and the growth traits of the adult fish fed on iced fish(P>0.05). But in the juvenile fish population fed on formulated feed, the T+156A SNP was significantly related to the full or empty of the stomach and intestine. And the fish with AA genotype of T+156A SNP have significantly higher growth trait values in body weight and body length than those with TT genotype(P<0.05). The results hinted that AA genotype in T+156A SNP has significant association with the adaptability of largemouth bass to the artificial diets. LPL type1 gene can be used as a candidate gene for the largemouth bass fed on formulated feeds, with high survival rate and fast growth rate.更多还原