豆状带绦虫线粒体rrnL和nad5基因的克隆及种系发育研究

›› 2013, Vol. 40 ›› Issue (6): 45-49.

豆状带绦虫线粒体rrnL和nad5基因的克隆及种系发育研究

范艺凡^1,2, 刘国华³, 杨言川⁴, 韩进欢⁴, 朱兴全¹, 蒋文灿², 孙晓林⁴

1. 中国农业科学院兰州兽医研究所, 家畜疫病病原生物学国家重点实验室, 甘肃兰州 730046;
2. 四川农业大学动物医学院, 四川雅安 625014;
3. 湖南农业大学动物医学院, 湖南长沙 410128;
4. 甘肃农业大学动物医学院, 甘肃兰州 730070

收稿日期:2012-11-19 出版日期:2013-06-20 发布日期:2013-06-20
通讯作者: 孙晓琳 E-mail:ndsxl@yahoo.com.cn
作者简介:范艺凡(1987-),女,四川人,硕士生,研究方向:寄生虫分子生物学。
基金资助:
全国农业科研杰出人才项目;家畜疫病病原生物学国家重点实验室开放基金(SKLVEB2010KFKT010、SKLVEB2011KFKT004)。

Cloning and Phylogenetic Analysis of Mitochondrial rrnL and nad5 Genes of Taenia pisiformis Isolates

FAN Yi-fan^1,2, LIU Guo-hua³, YANG Yan-chuan⁴, HAN Jin-huan⁴, ZHU Xing-quan¹, JIANG Wen-can², SUN Xiao-lin⁴

1. State Key Laboratory of Veterinary Etiological Biology, Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Lanzhou 730046, China;
2. College of Veterinary Medicine, Sichuan Agricultural University, Ya’an 625014 China;
3. College of Veterinary Medicine, Hunan Agricultural University, Changsha 410128, China;
4. College of Veterinary Medicine, Gansu Agricultural University, Lanzhou 730070, China

Received:2012-11-19 Online:2013-06-20 Published:2013-06-20

摘要/Abstract

摘要： 本研究旨在阐明中国豆状带绦虫囊尾蚴河南分离株线粒体核糖体大亚基基因(rrnL)部分序列(prrnL)和烟酰胺腺嘌呤二核苷酸(NADH)脱氢酶亚单位5基因(nad5)部分序列(pnad5)的遗传变异情况,并用prrnL和pnad5序列重构豆状带绦虫与其他带科绦虫的种系发育关系。采用聚合酶链式反应(PCR),扩增豆状带绦虫囊尾蚴分离株的线粒体prrnL和pnad5,将所测的序列用PAUP 4.0 Beta 10程序MP法绘制种系发育树,利用DNAStar 5.0中的MegAlign程序进行同源性分析。所获得的豆状带绦虫线粒体prrnL长度约为847 bp,pnad5为602 bp。所获rrnL序列与GenBank中豆状带绦虫相应序列的相似性为99.1%~100%,nad5为99.2%~100%;河南分离株之间rrnL序列的相似性为99.65%,nad5为99.5%;与GenBank中其他带科绦虫序列的相似度rrnL低于83.4%,nad5低于85.7%。种系发育分析结果表明,所有豆状带绦虫分离株和已知豆状带绦虫位于同一分支。由于豆状带绦虫分离株的线粒体rrnL和nad5基因序列种内很保守,种间差异较大,故可作为带科绦虫种间遗传变异研究的标记。

关键词: 豆状带绦虫; 线粒体DNA; rrnL基因; nad5基因; 种系发育关系

Abstract: The objectives of the present study were to examine sequence variation in the mitochondrial large subunit ribosomal rRNA (rrnL) and NADH dehydrogenase subunit 5 (nad5) genes among Taenia pisiformis isolates from Henan province and study its phylogenetic relationships with other cestodes using rrnL and nad5 sequences. The partial rrnL (prrnL) and nad5 (pnad5) genes sequences were amplified from each T.pisiformis sample by polymerase chain reaction (PCR), and prrnL and pnad5 sequences were aligned using the ClustalX 1.81. Maximum parsimony (MP) tree was constructed using the software PAUP 4.0 Beta 10. The lengths of prrnL and pnad5 sequences were 847 and 602 bp, respectively. Sequence similarity in rrnL among the examined T.pisiformis samples were 99.65%, and 99.5% for nad5, whereas inter-specific similarity among taeniid cestodes were significantly lower than 83.4% for rrnL, and lower than 85.7% for nad5. Phylogenetic analysis showed that all the T.pisiformis isolates clustered in the same clade. It was concluded that prrnL and pnad5 sequences could be used as a genetic marker for the identification and differentiation of taeniid cestodes.

Key words: Taenia pisiformis; mitochondrial DNA; rrnL gene; nad5 gene; phylogenetic relationship

中图分类号:

范艺凡, 刘国华, 杨言川, 韩进欢, 朱兴全, 蒋文灿, 孙晓林. 豆状带绦虫线粒体rrnL和nad5基因的克隆及种系发育研究[J]. , 2013, 40(6): 45-49.

FAN Yi-fan, LIU Guo-hua, YANG Yan-chuan, HAN Jin-huan, ZHU Xing-quan, JIANG Wen-can, SUN Xiao-lin. Cloning and Phylogenetic Analysis of Mitochondrial rrnL and nad5 Genes of Taenia pisiformis Isolates[J]. , 2013, 40(6): 45-49.

参考文献

1 Bogh H O, Zhu X Q, Qian B Z, et al. Scanning for nucleotide variations in mitochondrial DNA fragments of Schistosoma japonicum by single-strand conformation polymorphism[J]. Parasitology, 1999, 118(1): 73~82.
2 Boore J L.Animal mitochondrial genomes[J]. Nucleic Acids Res, 1999, 27(8): 1767~1780.
3 Gasser R B, Zhu X Q, Woods W. Genotyping Taenia tapeworms by singles-trand conformation polymorphism of mitochondrial DNA[J]. Electrophoresis, 1999, 20(14): 2834~2837.
4 Jia W Z, Yan H B, Guo A J, et al. Complete mitochondrial genomes of Taenia multiceps, T.hydatigena and T.pisiformis: Additional molecular markers for a tapeworm genus of human and animal health significance[J]. BMC Genomics, 2010, 11: 447.
5 Hu M,Chilton N B,Gasser R B. The mitochodrial genomics of parasitic nematodes of socio-economic importance:Recent progress,and implications for population genetics and systematics[J]. Adv Parasitol, 2004, 56: 133~212.
6 Liu G H, Lin R Q, Li M W, et al. The complete mitochondrial genomes of three cestode species of Taenia infecting animals and humans[J]. Mol Biol Rep, 2011, 38(4): 2249~2256.
7 Liu G H, Gasser R B, Su A, et al. Clear genetic distinctiveness between human- and pig-derived Trichuris based on analyses of mitochondrial datasets[J]. PLoS Negl Trop Dis, 2012, 6(2): e1539.
8 McManus D P, Bowles J. Molecular genetic approaches to parasite identification: Their value in diagnostic parasitology and systematics[J]. Int J Parasitol, 1996, 26(7): 687~704.
9 Pawlowski Z, Allan J, Sarti E, et al. Control of Taenia solium taeniasis/cysticercosis: From research towards implementation[J]. Int J Parasitol, 2005, 35(11~12): 1221~1232.
10 Sweeny J P, Ryan U M, Robertson I D, et al. Development of a modified molecular diagnostic procedure for the identification and quantification of naturally occurring strongylid larvae on pastures[J]. Vet Parasitol, 2012, 190(3~4): 467~481.
11 Toral-Bastida E, Garza-Rodriguez A, Jimenez-Gonzalez D E, et al. Development of Taenia pisiformis in golden hamster (Mesocricetus auratus)[J]. Parasit Vectors, 2011, 4: 147.
12 Yang D, Fu Y, Wu X, et al. Annotation of the transcriptome from Taenia pisiformis and its comparative analysis with three Taeniidae species[J]. PLoS One, 2012, 7(4): e32283.

[1]	李中波, 侯强红, 段德勇. 不同种群白纹伊蚊单倍型多态性、遗传分化及种系发育关系分析[J]. 中国畜牧兽医, 2023, 50(4): 1452-1460.
[2]	韩大勇, 周春宝, 倪黎纲, 陈章言, 赵锦明. 苏姜猪mtDNA D-loop序列的遗传多样性分析[J]. 中国畜牧兽医, 2023, 50(4): 1472-1479.
[3]	刘晓玮, 李瑞香, 王兴龙, 李晓. 基于mtDNA Cytb基因和D-loop区序列的陕西林麝遗传多样性分析[J]. 中国畜牧兽医, 2022, 49(4): 1352-1363.
[4]	李莹, 何国戈, 王艳, 何静怡, 黄爱珍, 郑经成, 葛影影, 罗成龙. 4个优质鸡群体mtDNA D-loop区遗传多样性分析[J]. 中国畜牧兽医, 2020, 47(2): 469-478.
[5]	卜李那, 彭业博, 张春媛, 刘洋秀, 赵毅强. 基于祖源分析的猪遗传成分鉴定[J]. 中国畜牧兽医, 2020, 47(2): 544-553.
[6]	李兴才, 潘成勇, 李辉, 杨华婷, 陈林, 肖涛. 香炉山鸡线粒体DNA D-Loop区遗传多样性及系统进化研究[J]. 中国畜牧兽医, 2019, 46(9): 2633-2641.
[7]	徐冰冰, 张家新, 王瑞军, 王志英, 李学武, 宋国欣, 张美娜, 郭富强, 张俊杰, 张燕军, 苏蕊, 刘志红, 李金泉. 哺乳动物精子线粒体研究进展[J]. 中国畜牧兽医, 2019, 46(9): 2651-2657.
[8]	李义书, 侯冠彧, 曹婷, 施力光, 周汉林, 周雄. 基于线粒体DNA D-loop区全序列分析儋州鸡遗传多样性及其起源进化关系[J]. 《中国畜牧兽医》, 2019, 46(5): 1439-1446.
[9]	胡慧艳, 贾青, 刘津, 侯胜奎, 赵思思, 张伟峰. 中国地方猪种线粒体基因组遗传多样性及其系统发育分析[J]. 《中国畜牧兽医》, 2017, 44(8): 2360-2368.
[10]	尹亚军, 杜伟立, 王令, 张涛. 略阳乌鸡线粒体DNA控制区(D-loop)的遗传多样性分析[J]. 《中国畜牧兽医》, 2017, 44(1): 173-179.
[11]	张冰, 廖海洪, 农素群, 马青艳, 孙俊丽. 广西5个猪种和国外3个猪种mtDNA D-loop序列遗传多样性及进化分析[J]. , 2015, 42(7): 1807-1815.
[12]	杨德英, 杨光友. 兔豆状囊尾蚴病研究进展[J]. , 2015, 42(4): 1015-1020.
[13]	王文涛，林家栋，陈眷华，张福平. 贵州地方鸡种的遗传变异研究[J]. 中国畜牧兽医, 2014, 41(4): 185-188.
[14]	王燕, 杨言川, 张悦文, 李佳缘, 朱兴全, 刘国华. 羊毛尾线虫线粒体nad4基因的序列变异分析[J]. , 2013, 40(4): 81-84.
[15]	宋国华, 陈朝阳, 庞文彪, 高继萍, 岳文斌. 中国地鼠线粒体D-loop基因的克隆及序列分析[J]. , 2013, 40(2): 126-129.