大口黑鲈病毒的定量聚合酶链反应检测

Quantitative polymerase chain reaction assay for largemouth bass virus.

作者信息

Getchell R G, Groocock G H, Schumacher V L, Grimmett S G, Wooster G A, Bowser P R

机构信息

Aquatic Animal Health Program, Department of Microbiology and Immunology, College of Veterinary Medicine, Cornell University, Ithaca, New York 14853, USA.

出版信息

J Aquat Anim Health. 2007 Dec;19(4):226-33. doi: 10.1577/H07-009.1.

DOI:10.1577/H07-009.1

PMID:18333479

Abstract

The use of quantitative polymerase chain reaction (QPCR) to test for largemouth bass virus (LMBV) was evaluated during a challenge experiment in which largemouth bass Micropterus salmoides were immersed in the type strain of LMBV. The real-time PCR and cell culture methods were both used to measure LMBV present in the inoculum. Additional samples tested by QPCR included gill, gonad, kidney, liver, mucus, spleen, and swim bladder. A plasmid clone containing a 248-base pair (bp) fragment of the major capsid protein gene (MCP*) was serially diluted and used as a standard to quantify the number of LMBV DNA copies present in the samples tested. A 62-bp fragment of DNA located in MCP* was amplified in the real-time PCR assay. This work has demonstrated the value of the QPCR assay in LMBV surveys.

摘要

在一项攻毒实验中，对大口黑鲈（Micropterus salmoides）进行了大口黑鲈病毒（LMBV）定量聚合酶链反应（QPCR）检测评估，实验中大口黑鲈被浸入LMBV的模式菌株中。实时PCR和细胞培养方法均用于测量接种物中存在的LMBV。通过QPCR检测的其他样本包括鳃、性腺、肾脏、肝脏、黏液、脾脏和鳔。含有主要衣壳蛋白基因（MCP*）248个碱基对（bp）片段的质粒克隆被连续稀释，并用作标准来量化测试样本中存在的LMBV DNA拷贝数。在实时PCR检测中扩增了位于MCP*中的一段62 bp的DNA片段。这项工作证明了QPCR检测在LMBV调查中的价值。