畜牧兽医学报2024，Vol.55Issue(5)：2080-2089,10.DOI:10.11843/j.issn.0366-6964.2024.05.026
基于Nanopore测序技术的非洲猪瘟病毒全基因组测序方法建立
A Whole Genome Sequencing Method for African Swine Fever Virus based on Nanopore Sequencing Technology was Established
周扬 1吴炜姿 1曹伟胜 2王福广 3许秀琼 3钟文霞 3吴立炀 3叶健 3卢受昇3
作者信息
- 1. 广东省动物疫病预防控制中心,广州 510230||华南农业大学兽医学院,广州 510642
- 2. 华南农业大学兽医学院,广州 510642
- 3. 广东省动物疫病预防控制中心,广州 510230
- 折叠
摘要
Abstract
African swine fever(ASF)is a highly contagious and deadly disease caused by the African swine fever virus(ASFV),which has dealt a heavy blow to the healthy development of China's pig industry in recent years.The large genome of ASFV makes it difficult to obtain whole-genome sequence about epidemic strains in a timely manner.This study aims to establish a simple and reliable ASFV whole-genome sequencing method using Nanopore third-generation sequencing technology.Thirty-one primer pairs covering the entire ASFV genome were designed and divided into 4 primer pools to amplify the sample.The amplified product was sequenced by Nanopore sequencing technology,and the relevant bioinformatics analysis methods were further optimized,and finally the ASFV whole-genome sequencing method was successfully established.Whole-genome sequence of ASFV with a total length of 189 416 bp was successfully obtained from an environmental swab sample by this method.Validation through first-generation sequencing has shown that the result of this method is 100％consistent with first-generation sequencing results in key genes and certain variant positions,including B646L,EP402R,E183L,MGF_360-12L,MGF_505-3R,and I177L gene.At the whole-genome level,the consistency between the result of this method and next-generation sequencing(NGS)result is 99.94％.In addition,the utilization of Nanopore sequencing technology in this study revealed a 56 bp repeat sequence insertion within the intergenic region flanked by the NP1450L and NP419L genes.This insertion was subsequently confirmed via first-generation sequencing techniques.Intriguingly,NGS methods failed to detect this distinct variant feature.This study successfully established an ASFV whole-genome sequencing method based on Nanopore technology.This method demonstrates excellent simplicity and reliability,providing an essential tool for the current prevention and control and molecular epidemiological research of ASF.关键词
Nanopore测序/非洲猪瘟病毒/生物信息学分析Key words
nanopore sequencing/African swine fever virus/bioinformatics analysis分类
农业科技引用本文复制引用
周扬,吴炜姿,曹伟胜,王福广,许秀琼,钟文霞,吴立炀,叶健,卢受昇..基于Nanopore测序技术的非洲猪瘟病毒全基因组测序方法建立[J].畜牧兽医学报,2024,55(5):2080-2089,10.
