遗传 ›› 2018, Vol. 40 ›› Issue (8): 620-631.doi: 10.16288/j.yczz.18-091

• 综述 • 上一篇    下一篇

不同单细胞全基因组扩增方法的比较及MALBAC在辅助生殖中的应用

姚雅馨1,喇永富2,狄冉1,刘秋月1,胡文萍1,王翔宇1,储明星1()   

  1. 1. 中国农业科学院北京畜牧兽医研究所,农业部动物遗传育种与繁殖重点实验室,北京 100193
    2. 甘肃农业大学动物科学技术学院,兰州 730070
  • 收稿日期:2018-04-07 修回日期:2018-06-09 出版日期:2018-08-16 发布日期:2018-07-02
  • 通讯作者: 储明星 E-mail:mxchu@263.net
  • 作者简介:姚雅馨,博士,助理研究员,研究方向:动物遗传育种。E-mail: yaoyaxin0602@126.com。|喇永富,博士研究生,专业方向:动物遗传育种。E-mail: 545042995@qq.com。姚雅馨和喇永富为共同第一作者。
  • 基金资助:
    国家自然科学基金项目(31772580);国家自然科学基金项目(31472078);国家肉羊产业技术体系专项(CARS-38);中央级公益性科研院所基本科研业务费专项(Y2017JC24);中国农业科学院科技创新工程(ASTIP-IAS13);农业科研杰出人才及其创新团队项目和国家万人计划科技创新领军人才项目资助

Comparison of different single cell whole genome amplification methods and MALBAC applications in assisted reproduction

Yao Yaxin1,La Yongfu2,Di Ran1,Liu Qiuyue1,Hu Wenping1,Wang Xiangyu1,Chu Mingxing1()   

  1. 1. Key Laboratory of Animal Genetics, Breeding and Reproduction of Ministry of Agriculture, Institute of Animal Science, Chinese Academy of Agricultural Sciences, Beijing 100193, China
    2. College of Animal Science and Technology, Gansu Agricultural University, Lanzhou 730070, China
  • Received:2018-04-07 Revised:2018-06-09 Online:2018-08-16 Published:2018-07-02
  • Contact: Chu Mingxing E-mail:mxchu@263.net
  • Supported by:
    Supported by the National Natural Science Foundation of China(31772580);Supported by the National Natural Science Foundation of China(31472078);Earmarked Fund for China Agriculture Research System(CARS-38);Central Public-interest Scientific Institution Basal Research Fund(Y2017JC24);Agricultural Science and Technology Innovation Program of China(ASTIP-IAS13);China Agricultural Scientific Research Outstanding Talents and Their Innovative Teams Program, China High-level Talents Special Support Plan Scientific and Technological Innovation Leading Talents Program

摘要:

单细胞全基因组扩增(whole genome amplification, WGA)是指在单细胞水平对全基因组进行扩增的新技术,其原理是将分离的单个细胞的微量全基因组DNA进行扩增,获得高覆盖率的完整的基因组后进行高通量测序,用于揭示细胞异质性。目前,WGA方法主要包括引物延伸预扩增(primer extension preamplification PCR, PEP-PCR)、简并寡核苷酸引物PCR (degenerate oligonucleotide primed PCR, DOP-PCR)、多重置换扩增(multiple displacement amplification, MDA)、多次退火环状循环扩增(multiple annealing and looping-based amplification cycles, MALBAC)等。本文对不同的单细胞WGA方法的原理及应用情况分别进行了阐述,并对其扩增效率进行评价和比较,包括基因组覆盖度、均一性、重现性、SNV (single-nucleotide variants)和CNV (copy number variants)检测力等。综合对比不同单细胞WGA方法后发现,MALBAC的扩增均一性最高、等位基因脱扣率最低、重现性最好,且对于CNV和SNV的检测效果最好。本文还阐述了MALBAC技术在人类单精子减数重组、非整倍体分析以及人类卵细胞基因组研究中的应用。

关键词: 单细胞全基因组扩增, MALBAC, DOP-PCR, MDA, 单细胞

Abstract:

Single-cell whole genome amplification (WGA) is a new technology, which can amplify small amounts of DNA from single cell and obtain the high coverage whole genome DNA template for revealing cell heterogeneity. Single cell WGA methods mainly include primer extension preamplification PCR (PEP-PCR), degenerate oligonucleotide primed PCR (DOP-PCR), multiple displacement amplification (MDA), and multiple annealing and looping-based amplification cycles (MALBAC). In this review, we describe the principles and applications of different single cell genome wide amplification, and we evaluate and compare their amplification efficiency, including the coverage of genome, homogeneity, reproducibility, and detection power of single-nucleotide variants (SNV) and copy number variants (CNV). The results show that MALBAC have the highest amplification homogeneity, the lowest allelic gene knockdown rate, the best reproducibility, and the best detection effect on CNV and SNV. We also describe the applications of MALBAC in human single sperm meiosis, aneuploidy analysis, and human oocyte genome research.

Key words: single cell whole genome amplification, MALBAC, DOP-PCR, MDA, single cell