鳜PEP、LPL基因单核苷酸多态性研究与鱼食性驯化相关分析
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摘要
本研究在转录组测序的基础上采用直接测序法(DS)和错配引物设计酶切技术(CRS-PCR),率先在国内外对影响鳜摄食的消化、代谢的关键调控基因胃蛋白酶(pepsinogen, PEP)、脂蛋白脂酶(lipoprotein, LPL)基因单核苷酸多态性(SNP)进行检测,并进一步对两个基因检测到的SNP位点的和鳜食性驯化表型性状相关性进行研究。采用CRS-PCR和RFLP技术对鳜胃蛋白酶基因第5外显子到第8外显子进行了单核苷酸多态性遗传多态性检测和分析,结果在第7外显子处共检测到2个SNP位点(C1T, C52T),均为同义突变。分别利用限制性内切酶Hhal和EcoRV对这两个外显子突变点进行基因分型,发现PEP基因第7外显子1位点和52位点酶切后分别产生2种基因型和3种等位基因。利用卡方检验分析PEP基因2个SNP位点基因型分布在鳜两种不同食性驯化表型群体中的分布情况,结果表明PEP基因2个SNP位点基因型分布与鳜食性驯化表型不具显著差异性(P>0.05)。将2个SNP位点不同基因型组合成5种双倍型,卡方检验表明双倍型Dip1(CTCC)与Dip5(TTTT)在两群体中存在显著差异(P<0.05)。采用DS技术对鳜LPL基因第5外显子到第8外显子进行单核苷酸多态性遗传多态性检测和分析,结果在该基因第7外显子共检测到3个SNP位点(A25T, G26T, C29G),其中A25T、C29G为错义突变。利用卡方检验分析LPL基因3个SNP位点基因型在鳜两种不同食性驯化表型群体中的分布情况,结果表明LPL基因3个SNP位点基因型分与对鳜食性驯化表型不具显著差异性(P>0.05)。将3个SNP位点不同基因型组合成5种双倍型,卡方检验表明双倍型Dip2(ATTTCC)在鳜两种食性驯化表型分布中存在显著性差异(P<0.05)。本研究所筛的选标记可应用于易驯食鳜的辅助选育,为今后鳜功能基因多态性的进一步研究和加快鳜食性驯化育种的遗传进展打下基础。
Pepsinogen (PEP) and liportein lipase (LPL) are critical enzymes in the dietary protein and lipid levels, digestibility and metabolism. In the present study, analyses of single nucleotide polymorphisms (SNPs) on PEP and LPL in Siniperca chuatsi based on population sequencing by direct sequencing (DS) and created restriction site PCR (CRS-PCR), was firstly carried out. in order to investigate the relationship of the two genes polymorphisms and phenotype with special feed habit domestication. using the methods of CRS-PCR and RFLP, polymorphisms in PEP ranging from exon 5 to exon 8 (introns included) were analyzed, and two SNP sites in exon 7 (C1T, C52T) were identified, both of which were synonymous mutations.the analysis between domesticated and undomesticated populations by Chi-square test among two SNP sites show not significantly different (P>0.05). Five diplotypes were constructed based on two SNP sites in the experiment population. Chi-square test indicated that there was significant association between diplotypel and diplotype5 in domesticated and undomesticated populations (P<0.05).
Using the methods of DS, polymorphisms in LPL ranging from exon 5 to exon 8 (introns included) were analyzed, and three SNP sites in exon 7 (A25T, G26T, C29G) were identified, two of which are non-synonymous mutations. the analysis between domesticated and undomesticated populations by Chi-square test among three SNP sites show not significantly different (P>0.05). Five diplotypes were constructed based on three SNP sites in the experiment population. Chi-square test indicated that there was significant association between diplotype2 in domesticated and undomesticated populations (P<0.05).
These genes can consider as genetic markers for feed habit domestication phenotype in Siniperca chuatsi. In addition, this study laid the foundation for developement of feeding domesticated cultivation of Siniperca chuatsi research in future.
Pepsinogen (PEP) and liportein lipase (LPL) are critical enzymes in the dietary protein and lipid levels, digestibility and metabolism. In the present study, analyses of single nucleotide polymorphisms (SNPs) on PEP and LPL in Siniperca chuatsi based on population sequencing by direct sequencing (DS) and created restriction site PCR (CRS-PCR), was firstly carried out. in order to investigate the relationship of the two genes polymorphisms and phenotype with special feed habit domestication. using the methods of CRS-PCR and RFLP, polymorphisms in PEP ranging from exon 5 to exon 8 (introns included) were analyzed, and two SNP sites in exon 7 (C1T, C52T) were identified, both of which were synonymous mutations.the analysis between domesticated and undomesticated populations by Chi-square test among two SNP sites show not significantly different (P>0.05). Five diplotypes were constructed based on two SNP sites in the experiment population. Chi-square test indicated that there was significant association between diplotypel and diplotype5 in domesticated and undomesticated populations (P<0.05).
Using the methods of DS, polymorphisms in LPL ranging from exon 5 to exon 8 (introns included) were analyzed, and three SNP sites in exon 7 (A25T, G26T, C29G) were identified, two of which are non-synonymous mutations. the analysis between domesticated and undomesticated populations by Chi-square test among three SNP sites show not significantly different (P>0.05). Five diplotypes were constructed based on three SNP sites in the experiment population. Chi-square test indicated that there was significant association between diplotype2 in domesticated and undomesticated populations (P<0.05).
These genes can consider as genetic markers for feed habit domestication phenotype in Siniperca chuatsi. In addition, this study laid the foundation for developement of feeding domesticated cultivation of Siniperca chuatsi research in future.
引文
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