安徽农业科学2017，Vol.45Issue(13)：135-140,6.
斑石鲷卵鞭虫病病原的分子鉴定与系统发育分析
Molecular Identification and Phylogenetic Analysis of a Pathogenic Dinoflagellate Isolated from Spotted Knifejaw (Oplegnathus puncatus)
摘要
Abstract
[Objective] A pathogenic dinoflagellate (the Bohai-1407 isolate) was isolated from diseased Oplegnathus puncatus.To perform molecular identification and phylogenetic analysis of the Bohai-1407 isolate.[Method] Four pair of specific PCR primers were used to amplify the ribosomal DNA (rDNA) fragments of the Bohai-1407 isolate.The rDNA fragments were cloned and sequenced.The sequences of cloned rDNA fragments were assembled and then analyzed by Blast.The systematic taxonomy of the Bohai-1407 isolate was analyzed by constructed phylogenetic trees based on the rDNA sequences of 10 dinoflagellates in family Blastodiniidae and the rDNA sequences of 19 isolates/clones in species Amyloodinium ocellatum.[Result] Four ribosomal DNA (rDNA) fragments of the Bohai-1407 isolate were amplified.Sequence analysis showed that the sequenced rDNA operon were 6 530 bp in length.The rDNA operon was composed of 74 bp of partial external transcribed spacer (ETS),1 813 bp of 18S (also called small subunit,SSU),352 bp of internal transcribed spacers 1 (ITS1),159 bp of5.8S,698 bp of internal transcribed spacers 2 (ITS2) and 3388 bp of 23S (also called large subunit,LSU),followed by a 46 bp of partial non-transcribed spacer (NTS) in the 3'end.The sequence identity between the Baohai-1407 isolate and the Ningde1412 strain ofA.ocellatum was as high as 99.5％.Based on the SSU sequence of 10 dinoflagellates in family Blastodiniidae,a phylogenetic tree was constructed and the Bohai-1407 isolate clustered with isolates ofA.ocellatum.Two phylogenetic trees based on the ITS1 and ITS2 sequences of 19 isolates/clones in species A.ocellatum were also constructed.The Bohai-1407 isolate always clustered with the Gulf of Mexico isolate FL_21 (DQ490260.1).[Conclusion] The pathogenic dinoflagellate (the Bohai-1407 isolate) isolated from diseased O.puncatus was A.ocellatum according to the molecular identification results of rDNA.The Bohai-1407 isolate was most closely related to the Gulf of Mexico isolate FL_21 ofA.ocellatum.关键词
斑石鲷/眼点淀粉卵涡鞭虫/核糖体DNA/分子鉴定/系统发育分析Key words
Oplegnathus puncatus/Amyloodinium ocellatum/rDNA/Molecular identification/Phylogenetic analysis分类
农业科技引用本文复制引用
付泉洁,范超,谢国驷,叶仕根,史成银..斑石鲷卵鞭虫病病原的分子鉴定与系统发育分析[J].安徽农业科学,2017,45(13):135-140,6.基金项目
中国水产科学研究院中央级公益性科研院所基本科研业务费专项(2016HY-ZD0505,2017HY-ZD0303). （2016HY-ZD0505,2017HY-ZD0303）
