单核细胞增生李斯特菌的荧光扩增片段长度多态性（fAFLP）分析

Fluorescent amplified fragment length polymorphism (fAFLP) analysis of Listeria monocytogenes.

作者信息

Amar Corinne

机构信息

Gastrointestinal Bacteria Reference Unit, Public Health England, 61 Colindale Avenue, London, NW9 5EQ, UK,

出版信息

Methods Mol Biol. 2014;1157:95-101. doi: 10.1007/978-1-4939-0703-8_8.

DOI:10.1007/978-1-4939-0703-8_8

PMID:24792551

Abstract

Fluorescent amplified fragment length polymorphism (fAFLP) is based on the selective PCR amplification of restriction fragments from a digest of total genomic DNA. Genomic DNA extracted from a purified bacterial isolate is completely digested with two endonucleases generating fragments which are ligated to specific double-stranded adaptors. The ligated fragments are then amplified by PCR using fluorescently labelled primers. Fluorescent amplified fragments are separated by size on an automated sequencer with a size standard. fAFLP is a rapid, highly reproducible technique which can be used to discriminate and subtype Listeria monocytogenes strains.

摘要

荧光扩增片段长度多态性（fAFLP）基于对总基因组DNA消化产生的限制性片段进行选择性PCR扩增。从纯化的细菌分离物中提取的基因组DNA用两种核酸内切酶完全消化，产生的片段与特定的双链接头连接。然后使用荧光标记的引物通过PCR扩增连接的片段。荧光扩增片段通过带有大小标准的自动测序仪按大小分离。fAFLP是一种快速、高度可重复的技术，可用于区分单核细胞增生李斯特菌菌株并对其进行亚型分类。