白斑综合征病毒DETECTR检测系统的建立OA北大核心CSTPCD
Establishment of DETECTR detection system for white spot syndrome virus
将重组酶介导等温扩增技术(RAA)与成簇的规律间隔短回文重复序列及其相关蛋白12a(CRISPR-Cas 12a)检测系统相结合,针对白斑综合征病毒(WSSV)建立快速、灵敏、特异的DNA内切酶靶向CRISPR反式报告(DETECTR)检测系统.针对WSSV保守基因设计RAA引物与crRNA,通过实验筛选最优组合和最佳反应条件,构建WSSVDETECTR检测系统.对该系统进行灵敏度与特异性测试,与WOAH推荐的Real-time PCR和NestedPCR检测方法比较.结果显示,WSSV DETECTR检测系统最低检出限为10 copies/μL,灵敏度与Real-time PCR、Nested PCR检测方法相当(均为3.34×101 copies/μL),与其他病原之间无交叉反应,与Real-time PCR、Nested PCR对临床样本的检测结果相一致.结果表明,建立的WSSV DETECTR检测系统具有快速简单、灵敏、特异等优点,为白斑综合征病毒的快速检测提供了新的方法.
To establish a rapid,sensitive and specific DNA endonuclease targeted CRISPR trans re-porter(DETECTR)detection system for white spot syndrome virus(WSSV)by combining recombinase-medi-ated amplification(RAA)with CRISPR-Cas12a.RAA primers and crRNA were designed for the conserved genes of WSSV,and the optimal combination and best reaction condition were screened experimentally to con-struct the WSSV DETECTR detection system.The sensitivity and specificity of the system were tested and compared with the real-time PCR and Nested PCR methods recommended by WOAH.The Results showed that the WSSV DETECTR system has a minimum detection limit of 10 copies/μL,which is the same as real-time PCR and Nested PCR(3.34×101 copies/μL),there was no cross-reactivity with other pathogens and no differ-ence in the test results from real-time PCR and Nested PCR for clinical samples.In conclusion,WSSV DE-TECTR detection system has the advantages of being rapid and simple,sensitive and specific,and pro-vides a new method for the rapid detection of white spot syndrome virus.
徐博文;罗宝正;周傲白雪;廖秀云;蒋晓霞;邵建宏;沙才华;黄海超;赵福振;陈轩
拱北海关技术中心,广东珠海 519015
畜牧业
重组酶介导等温扩增技术(RAA)CRISPR-Cas12a白斑综合征病毒快速检测
recombinase-mediated amplificationCRISPR-Cas12awhite sport syndrome virusrapid detection
《中国兽医科学》 2024 (008)
1056-1064 / 9
广东省重点领域研发计划(2022B1111030001)
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