摘要
为建立一种灵敏、特异的快速检测异育银鲫寄生洪湖碘泡虫(Myxobolus honghuensis)的方法,本研究根据洪湖碘泡虫ITS-5.8S r DNA基因序列筛选出一对特异性引物Mh F/R,建立PCR检测方法,对反应条件进行优化,并通过特异性试验、灵敏性试验与临床检测验证其可行性。结果显示,建立的PCR检测方法能特异性扩增洪湖碘泡虫相应的基因片段,长度为479 bp,而对试验中其他9种粘孢子虫的扩增结果均为阴性;最低能检测0.1 pg的虫体基因组DNA。通过临床样品检测,PCR方法比显微镜检测的检出率提高了19.5%。结果表明,该PCR方法特异、灵敏,适用于洪湖碘泡虫的快速检测。
To establish a sensitive,specific and rapid method for detection of Myxobolus honghuensis infecting allogynogenetic gibel carp Carassius auratus gibelio（ Bloch）,a single PCR method was developed with primers Mh F / R targeting on its ITS- 5. 8S r DNA gene. After optimizating the reaction condition and system,a series of experiments on specificity and sensitivity were conducted. Additionally,36 field samples were tested by the established PCR and microscopic examination. The results showed that the developed PCR method was specific to amplify a 479 bp fragment using genomic DNA from M. honghuensis with a detection limit of 0. 1 pg,and no cross- reactions with other 9 myxosporean species tested. The clinical detection rate was 19. 5% higher than that of microscopic examination. The results suggest that the present PCR method is an efficient approach for molecular detection of M. honghuensis because of its specificity and sensitivity.
作者
李丹
柳阳
翟艳花
顾泽茂
LI Dan LIU Yang ZHAI Yan-hua GU Ze-mao(Department of Aquatic Animal Medicine, College of Fisheries, Huazhong Agricultural University, Wuhan 430070,China Freshwater Aquaculture Collaborative Innovation Center of Hubei Province, Wuhan 430070, China Key Lab of Freshwater Animal Breeding, Ministry of Agriculture, Wuhan 430070, China)
出处
《淡水渔业》
CSCD
北大核心
2017年第1期12-16,共5页
Freshwater Fisheries
基金
国家自然科学基金项目(31572233)
