摘要
目的 以 3个遗传病为例 ,探讨长链 RT- PCR在基因突变分析中的应用。 方法 采用长链 RT- PCR扩增 AL D、ATP7B和 FV的 m RNA编码区。从凝胶中回收预期扩增产物 ,再以分段或全长方式进行二次 PCR。对二次 PCR产物进行直接序列测定 ,查找序列突变。 结果 在长链 RT- PCR中 ,三种 m RNA的编码区全长均得到扩增 ,预期扩增产物分别为 2 4 4 0 ,4 4 80 ,6 789bp。在二次 PCR中 ,分段扩增策略和全长扩增策略均可获得大量的特异性产物 ,后者均可直接用于序列测定。 结论 长链 RT- PCR对于长链 m RNA的序列分析是一个简便可靠的方法。
Objective\ To study the application of long\|distance RT\|PCR in mutational analysis of disease genes.\ Methods\ Long RT\|PCR was used to amplify the coding region of adrenoleukodystrophy(ALD), hepatolenticular degenration gene ATP7B and factor Ⅴ mRNA.\ Second\|round PCR was performed on the desired PCR products recovered from gel slices in full length or in segments, followed by direct sequencing to find the mutations.\ Results\ The full\|length coding region of all three mRNAs was amplified with sizes of 2440, 4480 and 6789 bp respectively.\ The second\|round PCR, whether in full length or in segments, could provide enough specific products for direct sequencing.\ Conclusion\ Long RT\|PCR was a simple and reliable method for mutational analysis of disease genes
出处
《福建医科大学学报》
2003年第3期256-259,共4页
Journal of Fujian Medical University
关键词
逆转录聚合酶链反应
基因
突变
肝豆状核变性
肾上腺脑白质营养不良
因子V
reverse transcriptase polymerase chain reaction
gene
mutation
hepatolenticular \{degenration\}
adrenoleukodystrophy
factor Ⅴ
