Mutations in cardiac troponin I （cTnI） gene were assessed based on gene-chip technology.Special probes were designed to fabricate the low-density gene-chip,which could detect the mutations in exons 3,5,7,and 8 of th...Mutations in cardiac troponin I （cTnI） gene were assessed based on gene-chip technology.Special probes were designed to fabricate the low-density gene-chip,which could detect the mutations in exons 3,5,7,and 8 of the cTnI gene simultaneously.For each exon,two oligonucleotide sequences labeled with fluorescein at the 5＇-end were designed,one （oligonucleotide Ⅰ） simulating the wild type and the other （oligonucleotide Ⅱ） simulating the mutant.Oligonucleotides Ⅰ and Ⅱ were mixed together to simulate the heterozygote.After optimizing the hybridization protocols,the fabricated gene-chip could detect the mutations in the exons of the cTnI gene with relative high sensitivity and specificity.The fully complementary probe gave a fluorescent signal almost 50% stronger than that of the one-base mismatched one,which is in accordance with the result from a theoretical estimate. An applicable special gene-chip is available to investigate and diagnose familial hypertrophic cardiomyopathy （FHCM） after further improvement.展开更多
Hypertrophic cardiomyopathy (HCM) is one of the diseases damaging people health most badly and some mutations of exons in cardiac troponin I (cTnI) gene are closely associated with family hypertrophic cardiomyopathy (...Hypertrophic cardiomyopathy (HCM) is one of the diseases damaging people health most badly and some mutations of exons in cardiac troponin I (cTnI) gene are closely associated with family hypertrophic cardiomyopathy (FHCM).A microarray was fabricated to screen mutations in exons 3,5,7,and 8 in cTnI gene.Primers were designed for the PCR (polymerase chain reaction) to amplify the target DNA fragments from fresh blood samples.In order to simplify the PCR process,multiplex PCR technology was investigated in detail.The concentration of Mg^(2+) played an important role in multiplex PCR process,a properly low concentration of Mg^(2+) submitted a better speciality of PCR products.The speciality was also favored when the annealing temperature was reasonably enhanced and 64℃is the optimal annealing temperature for the multiplex PCR systems.When applying the fabricated gene-chip to detect the target fragments from PCR mixture,the signal intensity sequence is in accordance with that from theoretic estimate.展开更多
研究旨在通过全基因组选择信号分析,挖掘宁蒗高原鸡品种重要的种质特性基因。研究使用“神农一号”地方鸡液相基因芯片对3个地方鸡品种(30只宁蒗高原鸡,30只正阳三黄鸡及33只固始鸡)进行基因分型。试验首先通过构建系统发育树、主成分...研究旨在通过全基因组选择信号分析,挖掘宁蒗高原鸡品种重要的种质特性基因。研究使用“神农一号”地方鸡液相基因芯片对3个地方鸡品种(30只宁蒗高原鸡,30只正阳三黄鸡及33只固始鸡)进行基因分型。试验首先通过构建系统发育树、主成分分析、祖先成分分析等方法,探究品种的种群遗传结构;然后,结合群体遗传分化指数(Pattern of genetic differentiation,FST)和跨群体单倍型统计(Cross-population haplotype-based statistic,XP-nSL)等比较基因组方法鉴定宁蒗高原鸡全基组选择信号;最后,对宁蒗高原鸡进行连续性纯合片段(Run of homozygosity,ROH)分析,以进一步探究其受到的人工和自然选择足迹。结果显示:选择信号分析发现宁蒗高原鸡在高海拔适应、骨骼和肌肉发育、生长及体型大小等性状经历了更高强度选择;ROH分析共鉴定到7个ROH岛,对其进行基因注释发现,与生长、体型大小、骨骼发育及高海拔适应等相关的基因受到了选择。研究表明,3个品种间有明显的群体分化,两个河南地方鸡品种遗传距离较近,而宁蒗高原鸡与两者明显分离,该结果与现实中分群结果一致,结果为宁蒗高原鸡遗传资源保护、开发利用与评价工作提供参考。展开更多
基金The National Natural Science Foundation of China(No.60071001)China Postdoctoral Science Foundation(No.2002)+2 种基金Trans-Century Training Programme Foundation for the Talents by theState Education Commission(No.[2004]527)the Foundation of the 135?Key Laboratory of Jiangsu Province(No.SK200205)the HighTechnology Research Plan of Jiangsu Province(No.BG2003033,BG2001010).
文摘Mutations in cardiac troponin I （cTnI） gene were assessed based on gene-chip technology.Special probes were designed to fabricate the low-density gene-chip,which could detect the mutations in exons 3,5,7,and 8 of the cTnI gene simultaneously.For each exon,two oligonucleotide sequences labeled with fluorescein at the 5＇-end were designed,one （oligonucleotide Ⅰ） simulating the wild type and the other （oligonucleotide Ⅱ） simulating the mutant.Oligonucleotides Ⅰ and Ⅱ were mixed together to simulate the heterozygote.After optimizing the hybridization protocols,the fabricated gene-chip could detect the mutations in the exons of the cTnI gene with relative high sensitivity and specificity.The fully complementary probe gave a fluorescent signal almost 50% stronger than that of the one-base mismatched one,which is in accordance with the result from a theoretical estimate. An applicable special gene-chip is available to investigate and diagnose familial hypertrophic cardiomyopathy （FHCM） after further improvement.
文摘Hypertrophic cardiomyopathy (HCM) is one of the diseases damaging people health most badly and some mutations of exons in cardiac troponin I (cTnI) gene are closely associated with family hypertrophic cardiomyopathy (FHCM).A microarray was fabricated to screen mutations in exons 3,5,7,and 8 in cTnI gene.Primers were designed for the PCR (polymerase chain reaction) to amplify the target DNA fragments from fresh blood samples.In order to simplify the PCR process,multiplex PCR technology was investigated in detail.The concentration of Mg^(2+) played an important role in multiplex PCR process,a properly low concentration of Mg^(2+) submitted a better speciality of PCR products.The speciality was also favored when the annealing temperature was reasonably enhanced and 64℃is the optimal annealing temperature for the multiplex PCR systems.When applying the fabricated gene-chip to detect the target fragments from PCR mixture,the signal intensity sequence is in accordance with that from theoretic estimate.
文摘研究旨在通过全基因组选择信号分析,挖掘宁蒗高原鸡品种重要的种质特性基因。研究使用“神农一号”地方鸡液相基因芯片对3个地方鸡品种(30只宁蒗高原鸡,30只正阳三黄鸡及33只固始鸡)进行基因分型。试验首先通过构建系统发育树、主成分分析、祖先成分分析等方法,探究品种的种群遗传结构;然后,结合群体遗传分化指数(Pattern of genetic differentiation,FST)和跨群体单倍型统计(Cross-population haplotype-based statistic,XP-nSL)等比较基因组方法鉴定宁蒗高原鸡全基组选择信号;最后,对宁蒗高原鸡进行连续性纯合片段(Run of homozygosity,ROH)分析,以进一步探究其受到的人工和自然选择足迹。结果显示:选择信号分析发现宁蒗高原鸡在高海拔适应、骨骼和肌肉发育、生长及体型大小等性状经历了更高强度选择;ROH分析共鉴定到7个ROH岛,对其进行基因注释发现,与生长、体型大小、骨骼发育及高海拔适应等相关的基因受到了选择。研究表明,3个品种间有明显的群体分化,两个河南地方鸡品种遗传距离较近,而宁蒗高原鸡与两者明显分离,该结果与现实中分群结果一致,结果为宁蒗高原鸡遗传资源保护、开发利用与评价工作提供参考。
