应用深度测序技术鉴定平菇球形病毒

doi:10.13560/j.cnki.biotech.bull.1985.2014.12.014

生物技术通报 ›› 2014, Vol. 0 ›› Issue (12): 86-91.doi: 10.13560/j.cnki.biotech.bull.1985.2014.12.014

应用深度测序技术鉴定平菇球形病毒

王少杰¹ ,王廿¹ ,师迎春² ,胡晓艳3, 周涛¹

1. 中国农业大学植物病理学系，北京 100193; 2. 北京市植物保护站，北京 100029; 3. 北京市农业技术推广站，北京 100029

收稿日期:2014-05-04 出版日期:2014-12-08 发布日期:2014-12-12
作者简介:王少杰，男，硕士研究生，研究方向:植物病毒;E-mail:wangshaojiestrong@163.com 通讯作者:周涛，男，副教授，研究方向:植物病毒;E-mail:taozhoucau@cau.edu.cn
基金资助:
2013 年食用菌创新团队岗位专家工作经费(PXM2013_036203_000055)

Identification of Oyster Mushroom Spherical Virus in Pleurotus ostreatus by Deep Sequencing

¹Wang Shaojie,¹Wang Nian,²Shi Yingchun,³Hu Xiaoyan,¹Zhou Tao

(1. College of Agriculture and Biotechnology, China Agriculture University, Beijing 100193; 2. Beijing Plant Protection Station, Beijing 100029; 3. Beijing Agricultural Technical Extension Station, Beijing 100029)

Received:2014-05-04 Published:2014-12-08 Online:2014-12-12

摘要/Abstract

摘要： 旨在鉴定采集于北京顺义的畸形平菇样品中的病毒，提取总 RNA，构建小RNA 库，对小RNA 库进行深度测序。测序结果表明，与平菇球形病毒(Oyster mushroom isometric virus，OMSV)基因组序列(NC_004560.1)匹配的小 RNA 共有 3075 条，分布在 OMSV 基因组的不同位置，形成一些热点区域。经拼接，获得了3条长度大于 500 个核苷酸的序列。根据其中覆盖 OMSV 外壳蛋白(Coat protein，CP)基因的序列(seq22)中相对保守区域，设计引物，利用 RT-PCR 扩增获得cp 基因部分序列。序列比对表明，其与 OMSV 相应区域序列相似度为 91%，证实了 OMSV 在畸形平菇样品中的侵染。同时，将深度测序结果与现有4种 siRNA 预测软件的预测结果进行比较，发现测序结果与软件预测结果之间存在差异，讨论了这一差异出现的原因。

关键词: 平菇, 深度测序, 平菇球形病毒, 小干扰RNA, 重叠群

Abstract: To identify viruses in malformed Pleurotus ostreatus samples collected in Shunyi district in Beijing, deep sequencing of small RNA from total RNA was performed. Consequently, there were 3 075 small interfering RNAs(siRNAs)pairing to the complete sequence of Oyster mushroom isometric virus(OMSV)(NC_004560.1). These siRNAs were located in most regions on genome of OMSV, some of which formed hot-spots. By putting these siRNAs together, three contigs with length more than 500 nucleotides were assembled. According to the sequence of one of these contigs, seq22, a pair of primers was designed to amplify partial coat protein gene via RT-PCR. The results showed that cloned sequence was similar to the reference sequence and to the contig with identities of 91% and 99%, respectively. These results confirmed OMSV in the collected mushroom samples. Meanwhile, the related data acquired from deep sequencing was compared to the predicted siRNAs obtained by four softwares, showing that they were different from each other. The difference was discussed.

Key words: Pleurotus ostreatus, Deep sequencing, Oyster mushroom spherical virus, siRNA, Contig

王少杰 ,王廿 ,师迎春 ,胡晓艳3, 周涛. 应用深度测序技术鉴定平菇球形病毒[J]. 生物技术通报, 2014, 0(12): 86-91.

Wang Shaojie,Wang Nian,Shi Yingchun,Hu Xiaoyan,Zhou Tao. Identification of Oyster Mushroom Spherical Virus in Pleurotus ostreatus by Deep Sequencing[J]. Biotechnology Bulletin, 2014, 0(12): 86-91.

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应用深度测序技术鉴定平菇球形病毒

Identification of Oyster Mushroom Spherical Virus in Pleurotus ostreatus by Deep Sequencing

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