摘要
基因全长序列是研究新基因功能的前提基础,基于PCR(polymerase chain reaction)建立了反向PCR、外源接头介导PCR和随机引物PCR三类基因序列全长扩增方法。虽然早期建立的反向PCR连接效率较低,易产生较高的非特异性扩增,但简单快捷的特点使其在鉴定T-DNA插入位点、基因融合位点等方面应用较多;外源接头介导PCR则因特异性的提高而备受欢迎,并衍生出单特异引物PCR、捕获PCR和步降PCR等多种技术,取得了较理想的扩增效果;而在大量样品分析中,自动化程度较高的随机引物PCR则更具优势。随着功能强大的聚合酶、连接酶、反转录酶的发现,推动了各种基于PCR的基因序列全长获取策略的发展,很多策略也因此而得以改进和整合,并获取更理想的实验结果,为研究基因结构、功能及其表达产物特性的研究提供了更完整的信息。
Full-length sequence is the basic premise of the study on genetic function of new gene, three amplification methods of the full-length sequence of gene, inverse PCR, adapter-mediated PCR and random primer PCR, have been established based on PCR （polymerase chain reaction）. Although the early established inverse PCR is inefficient in the ligation procedure and bring forth more non-specific amplification, it＇ s extensively used in the identification of T-DNA insertion sites and gene fusion sites for its simplicity and quickness. With the improvement of the specificity, adapter-mediated PCR is getting more and more popular, many novel technologies, such as single specific primer PCR, capture PCR and step-down PCR, were derived, and the ideal amplification effects were obtained. Recently, owing to its high degree of automation, random primer PCR is playing an important role in the analysis of a large number of samples. With the discovery of powerful enzymes, such as, DNA polymerase, ligase and reverse transcriptase, the development of the strategies for obtaining full-length sequence by PCR amplification technology were promoted, many strategies were improved and integrated, and better experimental results were obtained, it provided more complete information for the study of gene structure, function and transcript characteristics.
出处
《中国生物工程杂志》
CAS
CSCD
北大核心
2012年第11期115-123,共9页
China Biotechnology
基金
国家"十二五"科技支撑计划资助项目(2011BAD19B02)
关键词
全长序列
染色体步移RACE技术
Full-length sequence Chromosome walking RACE technology
