侧翼序列获取技术研究进展

doi:10.16288/j.yczz.21-415

遗传 ›› 2022, Vol. 44 ›› Issue (4): 313-321.doi: 10.16288/j.yczz.21-415

侧翼序列获取技术研究进展

徐纪明(), 朱建树, 李梦真, 胡晗, 毛传澡()

浙江大学生命科学学院植物生物学研究所,杭州 310058

收稿日期:2021-12-02 修回日期:2022-03-08 出版日期:2022-04-20 发布日期:2022-03-25
通讯作者: 毛传澡 E-mail:xujiming@zju.edu.cn;mcz@zju.edu.cn
作者简介:徐纪明,博士,助理研究员,研究方向：作物磷高效分子机制。E-mail: xujiming@zju.edu.cn
基金资助:
国家自然科学基金项目资助编号(32002121)

Progress on methods for acquiring flanking genomic sequence

Jiming Xu(), Jianshu Zhu, Mengzhen Li, Han Hu, Chuanzao Mao()

Institute of Plant Biology, College of Life Science, Zhejiang University, Hangzhou 310058, China

Received:2021-12-02 Revised:2022-03-08 Online:2022-04-20 Published:2022-03-25
Contact: Mao Chuanzao E-mail:xujiming@zju.edu.cn;mcz@zju.edu.cn
Supported by:
Supported by the National Natural Science Foundation of China No(32002121)

摘要/Abstract

摘要：

侧翼序列是指染色体中特定位点两侧的DNA序列,包含着候选基因、转录调控、染色体结构、生物安全等信息,在基因组学研究中具有重要的作用。侧翼序列获取技术主要应用于启动子和增强子等调控序列的克隆、鉴定T-DNA或转座子插入位点、染色体步移、全基因组空隙填补等,是结构基因组研究以及功能基因组研究的重要手段,在转基因动植物鉴定及安全管理等方面具有重要应用。随着分子生物学的发展,目前已经建立了许多侧翼序列的获取方法,依据技术原理可以分为质粒拯救法、反向PCR法、外源接头介导PCR法、半随机引物PCR法和基因组重测序法等5大类。本文系统总结了近年来侧翼序列获取技术的研究进展,并对这些技术的原理以及应用情况进行了较为系统的综述,为侧翼序列信息的获取提供参考。

关键词: 侧翼序列, 质粒拯救, 反向PCR, 外源接头PCR, 半随机引物PCR, 基因组重测序

Abstract:

Flanking genomic sequences refer to the DNA sequences flanking specific sites of known sequences in chromosome, which contain information such as candidate genes, transcriptional regulation, chromosome structure, and biosafety, and play an important role in genomics research. Flanking sequence acquisition technologies are mainly used in the cloning of regulatory sequences such as promoters and enhancers, identification of T-DNA or transposon insertion sites, chromosome walking, genome-wide gap filling, etc. It is an important means of structural genomics research and functional genomics research. It is applied in the identification of transgenic plants and animals and their safety management. With the development of molecular biology, many methods for obtaining flanking sequences have been established, including plasmid rescue, inverse PCR, ligation-mediated PCR, semi-random primer PCR, whole-genome resequencing etc. In this review, we summarize and compared different methods for acquiring flanking genomic sequence. The principles and research progress of each approach are discussed.

Key words: flanking sequence, plasmid rescue, inverse PCR, ligation-mediated PCR, semi-random primer PCR, whole-genome resequencing

徐纪明, 朱建树, 李梦真, 胡晗, 毛传澡. 侧翼序列获取技术研究进展[J]. 遗传, 2022, 44(4): 313-321.

Jiming Xu, Jianshu Zhu, Mengzhen Li, Han Hu, Chuanzao Mao. Progress on methods for acquiring flanking genomic sequence[J]. Hereditas(Beijing), 2022, 44(4): 313-321.

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方法	DNA酶切	DNA用量	限制因素	非特异性产物	灵敏度	操作复杂性
质粒拯救	需要	2~5 μg	酶切位点选择;连接或环化效率	菌落假阳性	低	复杂
反向PCR	需要	2~5 μg	酶切位点选择;连接或环化效率	非特异性连接	低	复杂
接头介导PCR	需要	2~5 μg	酶切位点选择;连接或环化效率	单引物扩增	中等	复杂
半随机引物PCR	不需要	20~200 ng	简并引物设计;插入载体完整性	单引物扩增	高	中等
基因组重测序	不需要	2~5 μg	测序深度;插入位置	无	非常高	简单

侧翼序列获取技术研究进展

Progress on methods for acquiring flanking genomic sequence

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