The information of single nucleotide polymorphisms （SNPs） is quite unknown in sweetpotato. In this study, two sweetpotato varieties （Xushu 18 and Xu 781） were sequenced by Illumina technology, as well as de novo t...The information of single nucleotide polymorphisms （SNPs） is quite unknown in sweetpotato. In this study, two sweetpotato varieties （Xushu 18 and Xu 781） were sequenced by Illumina technology, as well as de novo transcriptome assembly, functional annotation, and in silico discovery of potential SNP molecular markers. Tetra-primer Amplification Refractory Mutation System PCR （ARMS-PCR） is a simple and sufficient method for detecting different alleles in SNP locus. Total 153 sets of ARMS-PCR primers were designed to validate the putative SNPs from sequences. PCR products from 103 sets of primers were different between Xu 781 and Xushu 18 via agarose gel electrophoresis, and the detection rate was 67.32%. We obtained the expected results from 32 sets of primers between the two genotypes. Furthermore, we ascertained the optimal annealing temperature of 32 sets of primers. These SNPs might be used in genotyping, QTL mapping, or marker-assisted trait selection further in sweetpotato. To our knowledge, this work was the first study to develop SNP markers in sweetpotato by using tetra-primer ARMS-PCR technique. This method was a simple, rapid, and useful techn!que to develop SNP markers, and will provide a potential and preliminary application in discriminating cultivars in sweetpotato.展开更多
文摘【目的】早发性肌无力综合征(early onset muscle weakness syndrome,MW)是新近发现的一种荷斯坦牛遗传缺陷,患病犊牛表现为出生后趴卧不起、后肢肌肉萎缩等,其遗传机制与L型钙通道蛋白α1S亚基编码基因CACNA1S的单碱基突变相关。本研究旨在建立该遗传缺陷的分子检测方法,并探究其在国内荷斯坦牛群体中的扩散情况。【方法】基于MW致病位点特异性DNA序列,设计扩增阻滞突变系统-聚合酶链式反应(ARMS-PCR)检测引物,对317份荷斯坦牛冻精和毛囊样本进行MW遗传缺陷基因筛查,通过Sanger测序对ARMS-PCR检测结果进行验证,并利用系谱数据追溯MW突变源头。【结果】ARMS-PCR检出MW携带者21头,携带率为6.62%(21/317),未发现缺陷基因纯合子,提示该突变为隐性纯合致死。Sanger测序与ARMS-PCR所得基因型完全一致,证明该技术具有高度准确性。系谱追溯表明,MW携带者的遗传来源可追溯至1984年出生的荷斯坦公牛Southwind Bell of Bar-Lee。【结论】MW遗传缺陷已在中国荷斯坦牛群体中扩散且携带率较高,建议牧场应尽早开展MW遗传缺陷基因检测和风险评估,采取科学的选种选配措施以减少遗传缺陷导致的经济损失。
基金supported by the China Agriculture Research System (CARS-11)the National High-Tech R&D Program of China (2012AA101204)the Jiangsu Independent Inno vation Funds of Agriculture,China (CX(13)2032)
文摘The information of single nucleotide polymorphisms （SNPs） is quite unknown in sweetpotato. In this study, two sweetpotato varieties （Xushu 18 and Xu 781） were sequenced by Illumina technology, as well as de novo transcriptome assembly, functional annotation, and in silico discovery of potential SNP molecular markers. Tetra-primer Amplification Refractory Mutation System PCR （ARMS-PCR） is a simple and sufficient method for detecting different alleles in SNP locus. Total 153 sets of ARMS-PCR primers were designed to validate the putative SNPs from sequences. PCR products from 103 sets of primers were different between Xu 781 and Xushu 18 via agarose gel electrophoresis, and the detection rate was 67.32%. We obtained the expected results from 32 sets of primers between the two genotypes. Furthermore, we ascertained the optimal annealing temperature of 32 sets of primers. These SNPs might be used in genotyping, QTL mapping, or marker-assisted trait selection further in sweetpotato. To our knowledge, this work was the first study to develop SNP markers in sweetpotato by using tetra-primer ARMS-PCR technique. This method was a simple, rapid, and useful techn!que to develop SNP markers, and will provide a potential and preliminary application in discriminating cultivars in sweetpotato.
