染色体步移技术克隆已知序列侧翼启动子的研究进展.docVIP

Online system: 农 业 生 物 技 术 学 报 Journal of Agricultural Biotechnology 2013, 21(3): 355~366 DOI: 10.3969/j.issn.1674-7968.2013.03.013 评述与展望 Review and Progress 染色体步移技术克隆已知序列侧翼启动子的研究进展 康丹 1 方小艳 1 游腾飞 1 王婷婷 1 眭安平 2* 杨星勇 1，3* 1 西南大学生命科学学院，重庆 400715；2 西南大学图书馆，重庆 400715；3 重庆师范大学生命科学学院，重庆 401331 * 通讯作者，tobpol3@；yangxy94@  摘 要  启动子是基因表达调控的关键顺式作用元件，也是基因工程表达载体的一个重要元件。对启动 子结构和功能的阐释，有助于基因表达的时空控制。选择恰当方法克隆序列未知的启动子有着非常重要 的意义和广阔的应用前景。随着 PCR 技术的问世，基于 PCR 扩增的染色体步移技术进行启动子克隆具 有较好的可操作性，其在基因的分子生物学和基因工程研究方面也是一项重要的技术策略。本文讨论了 近年来以 PCR 为基础的染色体步移技术克隆启动子的方法，主要包括反向 PCR、接头介导 PCR、半随机 引物 PCR 三大类，归纳比较了其克隆原理和研究进展，为克隆已知序列旁侧未知启动子序列提供参考。 关键词 染色体步移，启动子，反向 PCR，接头介导 PCR，半随机引物 PCR  Progress of Chromosome Walking the Unknown Promoter Sequences Flanked by a Known Segment KANG Dan1 FANG Xiao-Yan1 YOU Teng-Fei1 WANG Ting-Ting1 SUI An-Ping2* YANG Xing-Yong1,3* 1 The School of Life Science, Southwest University,Chongqing 400715, China; 2 The Libarary of Southwest University, Chongqing 400715, China; 3 The College of Life Science, Chongqing Normal University, Chongqing 401331, China * Corresponding author, tobpol3@; yangxy94@ Abstract Promoter is a key cis-acting element to regulate gene e xpression, which is also an important element of the expression vector in genetic engineering. Understanding the structure and function of promoters contributes to constructing vector and controlling the gene expression, further we can achieve the goal that induces protein expression selectively. Therefore introducing a proper method to clone the promoter presents significance and wide application prospects. Since the invention of polymerase chain reaction (PCR) technique, several PCR-based chromosome walking methods have been developed to amplify unknown promoter sequences flanked by a known segment. In this review, we introduce the PCR-based methodologies for the rapid acquisition of unknown promoter, including Inverse PCR, Ligation-mediated PCR, Semi-random primer PCR. These principles and research progresses of ea