长沙地区7种寄主蚜虫的快速鉴定及其亲缘关系分析
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摘要
以线粒体基因COI片段为基础,分别通过DNA条形码及PCR-RFLP(经Dra I,Hinf I,Ssp I,Taq I 4种限制性内切酶酶切)两种方法对长沙湖南农业大学地区7种寄主上的蚜虫进行了快速鉴定。两种鉴定方法结果均表明,7种寄主上的蚜虫分属豆蚜(Aphis craccivora)、桃粉大尾蚜(Hyalopterus pruni)、棉蚜(Aphis gossypii)、蚊母新胸蚜(Neothoracaphis yanonis)、绣线菊蚜(Aphis spiraecola)及莴苣指管蚜(Uroleucon formosanum)6种。其中,豆蚜、棉蚜、绣线菊蚜推测为同一个属,与传统分类结果一致。通过对比发现,在蚜虫分类鉴定上,PCR-RFLP技术不仅具有与DNA条形码相同的准确性,且有更加经济快速的优点。
Based on the mitochondrial gene COI, two molecular identification methods, DNA barcoding and PCR-RFLP(four kinds of restriction endonuclease: Dra I, Hinf I, Ssp I, Taq I), were used to rapidly identify aphids on seven kinds of host plants in Changsha area. Aphids on seven hosts were identified respectively as Aphis craccivora, Hyalopterus pruni,A. gossypii, Neothoracaphis yanonis, A. spiraecola and Uroleucon formosanum, by both methods. It indicated that A.craccivora, A. gossypii and A. spiraecola belonged to the same genus, which were identical as the results by traditional classification methods. It was proved that PCR-RFLP technology was as accurate as DNA barcoding, while PCR-RFLP was more economical and rapid.
Based on the mitochondrial gene COI, two molecular identification methods, DNA barcoding and PCR-RFLP(four kinds of restriction endonuclease: Dra I, Hinf I, Ssp I, Taq I), were used to rapidly identify aphids on seven kinds of host plants in Changsha area. Aphids on seven hosts were identified respectively as Aphis craccivora, Hyalopterus pruni,A. gossypii, Neothoracaphis yanonis, A. spiraecola and Uroleucon formosanum, by both methods. It indicated that A.craccivora, A. gossypii and A. spiraecola belonged to the same genus, which were identical as the results by traditional classification methods. It was proved that PCR-RFLP technology was as accurate as DNA barcoding, while PCR-RFLP was more economical and rapid.
引文
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[5]陈倩,沈佐锐,王永模.蚜虫的表型可塑性及其遗传基础[J].昆虫学报,2006(5):859-66.
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[14]Sunnucks P,EnglanD P R,Taylor A C,et al.Microsatellite and chromosome Evolution of parthenogenetic sitobion aphids in Australia[J].Genetics,1996,144(2):747.
[15]杨效文,张广学,陈晓峰.棉蚜微卫星DNA的克隆及其多态性检测[J].昆虫学报,2001,44(4):586-589.
[16]张旭,金道超,郭建军,等.螨类系统学研究中的分子标记[J].昆虫知识,2008,(2):198-203.
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[20]Hebert P D N,Stoeckle M Y,Zemlak T S,et al.Identification of Birds through DNA Barcodes[J].Plos Biology,2004,2(10):e312.
[21]Meier R,Shiyang K,Vaidya G,et al.DNA barcoding andtaxonomy in Diptera:A tale of high intraspecific variability and low identification success[J].Systematic Biology,2006,55(5):715-728.
[22]Ratnasingham S,Hebert P D N.BOLD:The barcode of life data system(www.barcodinglife.org)[J].Molecular Ecology Notes,2007,7(3):355-364.
[23]温娟,陈睿,姜立云,等.基于DNA条形码对北京地区蔷薇科花卉上蚜虫的快速鉴定[J].应用昆虫学报,2013(1):29-40.
[24]宋韶彬,石志勇,金倩,等.DNA条形码技术在河北保定、廊坊地区鳞翅目昆虫上的应用及小型区域数据库的构建[J].应用昆虫学报,2014,(1):156-168.
[25]Song C,Lin X L,Wang Q,et al.DNA barcodes successfully delimit morphospecies in a superdiverse insect genus[J].Zoologica Scripta,2018,47(3):311-324.
[26]Hebert P D N,Ratnasingham S,Dewaard J R.Barcoding animal life:cytochrome c oxidase subunit 1 divergences among closely related species[J].Proceedings Biological Sciences,2003,270(S1):S96-99.
[27]Clark T L,Meinke L J,Foster J E.PCR-RFLP of the mitochondrial cytochrome oxidase(subunit I)gene provides diagnostic markers for selected Diabrotica species(Coleoptera:Chrysomelidae)[J].Bulletin of Entomological Research,2001,91(6):419-427.
[28]Fanello C,Santolamazza F,DELLA T A.Simultaneous identification of species and molecular forms of the Anopheles gambiae complex by PCR-RFLP[J].Medical&Veterinary Entomology,2002,16(4):461-464.
[29]Bosco D,Loria A,Sartor C,et al.PCR-RFLP identification of Bemisia tabaci biotypes in the Mediterranean Basin[J].Phytoparasitica,2006,34(3):243-251.
[30]Valenzuela I,Hoffmann A A,Malipatil M B,et al.Identification of aphid species(Hemiptera:Aphididae:Aphidinae)using a rapid polymerase chain reaction restriction fragment length polymorphism method based on the cytochrome oxidase subunit I gene[J].Austral Entomology,2007,46(4):305-312.
[31]林丽莉,吴佳教,曾玲,等.二种大实蝇的PCR-RFLP快速鉴定方法[J].昆虫知识,2007(4):588-592.
[32]Patrelle C,Ohst T,Picard D,et al.A new PCR-RFLP-based method for an easier systematic affiliation of European water frogs[J].Molecular Ecology Resources,2011,11(1):200-205.
[33]张龙霞,李惠萍,裴雁曦.微流控芯片技术用于三种小蠹COI-COII区的PCR-RFLP分析[J].应用昆虫学报,2012(6):1721-1725.
[34]Brunner P C,Fleming C,Frey J E.A molecular identification key for economically important thrips species(Thysanoptera:Thripidae)using direct sequencing and a PCR-RFLP-based approach[J].Agricultural&Forest Entomology,2015,4(2):127-136.
[35]Carreira G P,Mckeown N J,Shaw P W,et al.A PCR-RFLP method for stock assignment in the endemic Macaronesian gastropod,Patella candei d'Orbigny,and its potential for future conservation strategies[J].Fisheries Research,2018,199.
[36]张广学,陈小琳,乔格侠,等.西北农林蚜虫志:昆虫纲同翅目蚜虫类[M].北京:中国环境科学出版社,1999:1-563.
[37]郭昆.中国扁蚜科的系统分类研究(同翅目:蚜总科)[D].西安:陕西师范大学,2005.
[38]乔格侠,张广学,姜立云,等.河北动物志蚜虫类[M].石家庄:河北科学技术出版社,2009:1-622.
[39]Deng G R.A sensitive non-radioactive PCR-RFLP analysis for detecting point mutations at 12th codon of oncogene c-Ha-ras in DNAs of gastric cancer[J].Nucleic Acids Research,1988,16(13):6231.
[40]Hidetoski I.PCR-RFLP method holds great promise for complete HLA class II genotyping[J].Tissue Antigens,1990,36(2):88-92.
[41]Ota M,Seki T,Nomura N,et al.Modified PCR-RFLPmethod for HLA-DPB1 and-DQA1 genotyping[J].Tissue Antigens,1991,38(3):60-71.
[42]Eijk M J T V,Stewart-Haynes J A,Lewin H A.Extensive polymorphism of the BOLA-DRB3 gene distinguished by PCR-RFLP[J].Animal Genetics,1992,23(6):483-496.
[43]Navarro E,Simonet P,Normand P,et al.Characterization of natural populations of Nitrobacter spp.using PCR/RFLP analysis of the ribosomal intergenic spacer[J].Archives of Microbiology,1992,157(2):107.
[44]Tsukamoto K,Watanabe T,Matsushima T,et al.Determination by PCR-RFLP of apo E genotype in a Japanese population[J].Journal of Laboratory&Clinical Medicine,1993,121(4):598-602.
[45]Ponsonnet C,Nesme X.Identification of Agrobacterium strains by PCR-RFLP analysis of pTi and chromosomal regions[J].Archives of Microbiology,1994,161(4):300-309.
[46]Barth D,Krenek S,Fokin S I,et al.Intraspecific genetic variation in Paramecium revealed by mitochondrial cytochrome C oxidase I sequences[J].Journal of Eukaryotic Microbiology,2006,53(1):20-25.
[47]Webster B L,Emery A M,Webster J P,et al.Genetic Diversity within Schistosoma haematobium:DNA Barcoding Reveals Two Distinct Groups[J].Plos Neglected Tropical Diseases,2012,6(10):10.
[2]Qiao G X,Zhang G X,Simon J C,et al.Preliminary study of aphid diversity in China:taxonomic and geographic variation//Rispe C,HulléM,Dedryver C A,et al.Aphids in a new millennium[C].Paris:Institute National de la RecherchéAgronomique.2004:139-146.
[3]张利军,曹挥,李友莲.蚜虫分类研究综述[J].山西农业大学学报(自然科学版),2000(4):360-3.
[4]Agarwala B K.Phenotypic plasticity in aphids(Homoptera:Insecta):components of variation and causative factors[J].Current Science,2007,93(3):308-13.
[5]陈倩,沈佐锐,王永模.蚜虫的表型可塑性及其遗传基础[J].昆虫学报,2006(5):859-66.
[6]Gorur G,Lomonaco C,Mackenzie A.Phenotypic plasticity in host choice behavior in black bean aphid,Aphis fabae(Homoptera:Aphididae)[J].Arthropod-Plant Interactions,2007,1(3):187-94.
[7]赵冲,王秀芳,陈丹,等.不同地理种群烟蚜形态特征差异分析[J].昆虫学报,2013,56(12):1452-1463.
[8]张广学,乔格侠,钟铁森,等.中国动物志:昆虫纲.第14卷,同翅目纩蚜科瘿绵蚜科[M].北京:科学出版社,1999:1-380.
[9]Black W C,Duteau N M,Puterka G J,et al.Use of the random amplified polymorphic DNA polymerase chain reaction(RAPD-PCR)to detect DNA polymorphisms in aphids(Homoptera:Aphididae)[J].Bulletin of Entomological Research,1992,82(2):151-159.
[10]Kambhampati S,Black W C,Rai K S.Random amplified polymorphic DNA of mosquito species and populations(Diptera:Culicidae):techniques,statistical analysis,and applications[J].Journal of Medical Entomology,1992,29(6):939-945.
[11]Blanchetot A,Gooding R H.Genetic analysis by DNAfingerprinting in tsetse fly genomes[J].Insect Biochemistry and Molecular Biology,1993,23(8):937-944.
[12]Black W C.Variation in the ribosomal RNA cistron among host-adapted races of an aphid(Schizaphis graminum)[J].Insect Molecular Biology,1993,2(2):59-69.
[13]Osakabe M,Sakagami Y.RFLP analysis of ribosomal DNA in sibling species of spider mite,genus Panonychus(Acari:Tetranychidae)[J].Insect Molecular Biology,1994,3(1):63-66.
[14]Sunnucks P,EnglanD P R,Taylor A C,et al.Microsatellite and chromosome Evolution of parthenogenetic sitobion aphids in Australia[J].Genetics,1996,144(2):747.
[15]杨效文,张广学,陈晓峰.棉蚜微卫星DNA的克隆及其多态性检测[J].昆虫学报,2001,44(4):586-589.
[16]张旭,金道超,郭建军,等.螨类系统学研究中的分子标记[J].昆虫知识,2008,(2):198-203.
[17]Tautz D Arctander P,Minelli A,et al.DNA points the way ahead in taxonomy[J].Nature,2002,418(6897):479.
[18]Hebert P D N,Cywinska A,Ball S L,et al.Biological identifications through DNA barcodes[J].Proceedings Biological Sciences,2003,270(1512):313-321.
[19]Hebert P D N,Penton E H,Burns J M,et al.Ten species in one:DNA barcoding reveals cryptic species in the neotropical skipper butterfly Astraptes fulgerator[J].Proceedings of the National Academy of Sciences of the United States of America,2004,101(41):14812-14817.
[20]Hebert P D N,Stoeckle M Y,Zemlak T S,et al.Identification of Birds through DNA Barcodes[J].Plos Biology,2004,2(10):e312.
[21]Meier R,Shiyang K,Vaidya G,et al.DNA barcoding andtaxonomy in Diptera:A tale of high intraspecific variability and low identification success[J].Systematic Biology,2006,55(5):715-728.
[22]Ratnasingham S,Hebert P D N.BOLD:The barcode of life data system(www.barcodinglife.org)[J].Molecular Ecology Notes,2007,7(3):355-364.
[23]温娟,陈睿,姜立云,等.基于DNA条形码对北京地区蔷薇科花卉上蚜虫的快速鉴定[J].应用昆虫学报,2013(1):29-40.
[24]宋韶彬,石志勇,金倩,等.DNA条形码技术在河北保定、廊坊地区鳞翅目昆虫上的应用及小型区域数据库的构建[J].应用昆虫学报,2014,(1):156-168.
[25]Song C,Lin X L,Wang Q,et al.DNA barcodes successfully delimit morphospecies in a superdiverse insect genus[J].Zoologica Scripta,2018,47(3):311-324.
[26]Hebert P D N,Ratnasingham S,Dewaard J R.Barcoding animal life:cytochrome c oxidase subunit 1 divergences among closely related species[J].Proceedings Biological Sciences,2003,270(S1):S96-99.
[27]Clark T L,Meinke L J,Foster J E.PCR-RFLP of the mitochondrial cytochrome oxidase(subunit I)gene provides diagnostic markers for selected Diabrotica species(Coleoptera:Chrysomelidae)[J].Bulletin of Entomological Research,2001,91(6):419-427.
[28]Fanello C,Santolamazza F,DELLA T A.Simultaneous identification of species and molecular forms of the Anopheles gambiae complex by PCR-RFLP[J].Medical&Veterinary Entomology,2002,16(4):461-464.
[29]Bosco D,Loria A,Sartor C,et al.PCR-RFLP identification of Bemisia tabaci biotypes in the Mediterranean Basin[J].Phytoparasitica,2006,34(3):243-251.
[30]Valenzuela I,Hoffmann A A,Malipatil M B,et al.Identification of aphid species(Hemiptera:Aphididae:Aphidinae)using a rapid polymerase chain reaction restriction fragment length polymorphism method based on the cytochrome oxidase subunit I gene[J].Austral Entomology,2007,46(4):305-312.
[31]林丽莉,吴佳教,曾玲,等.二种大实蝇的PCR-RFLP快速鉴定方法[J].昆虫知识,2007(4):588-592.
[32]Patrelle C,Ohst T,Picard D,et al.A new PCR-RFLP-based method for an easier systematic affiliation of European water frogs[J].Molecular Ecology Resources,2011,11(1):200-205.
[33]张龙霞,李惠萍,裴雁曦.微流控芯片技术用于三种小蠹COI-COII区的PCR-RFLP分析[J].应用昆虫学报,2012(6):1721-1725.
[34]Brunner P C,Fleming C,Frey J E.A molecular identification key for economically important thrips species(Thysanoptera:Thripidae)using direct sequencing and a PCR-RFLP-based approach[J].Agricultural&Forest Entomology,2015,4(2):127-136.
[35]Carreira G P,Mckeown N J,Shaw P W,et al.A PCR-RFLP method for stock assignment in the endemic Macaronesian gastropod,Patella candei d'Orbigny,and its potential for future conservation strategies[J].Fisheries Research,2018,199.
[36]张广学,陈小琳,乔格侠,等.西北农林蚜虫志:昆虫纲同翅目蚜虫类[M].北京:中国环境科学出版社,1999:1-563.
[37]郭昆.中国扁蚜科的系统分类研究(同翅目:蚜总科)[D].西安:陕西师范大学,2005.
[38]乔格侠,张广学,姜立云,等.河北动物志蚜虫类[M].石家庄:河北科学技术出版社,2009:1-622.
[39]Deng G R.A sensitive non-radioactive PCR-RFLP analysis for detecting point mutations at 12th codon of oncogene c-Ha-ras in DNAs of gastric cancer[J].Nucleic Acids Research,1988,16(13):6231.
[40]Hidetoski I.PCR-RFLP method holds great promise for complete HLA class II genotyping[J].Tissue Antigens,1990,36(2):88-92.
[41]Ota M,Seki T,Nomura N,et al.Modified PCR-RFLPmethod for HLA-DPB1 and-DQA1 genotyping[J].Tissue Antigens,1991,38(3):60-71.
[42]Eijk M J T V,Stewart-Haynes J A,Lewin H A.Extensive polymorphism of the BOLA-DRB3 gene distinguished by PCR-RFLP[J].Animal Genetics,1992,23(6):483-496.
[43]Navarro E,Simonet P,Normand P,et al.Characterization of natural populations of Nitrobacter spp.using PCR/RFLP analysis of the ribosomal intergenic spacer[J].Archives of Microbiology,1992,157(2):107.
[44]Tsukamoto K,Watanabe T,Matsushima T,et al.Determination by PCR-RFLP of apo E genotype in a Japanese population[J].Journal of Laboratory&Clinical Medicine,1993,121(4):598-602.
[45]Ponsonnet C,Nesme X.Identification of Agrobacterium strains by PCR-RFLP analysis of pTi and chromosomal regions[J].Archives of Microbiology,1994,161(4):300-309.
[46]Barth D,Krenek S,Fokin S I,et al.Intraspecific genetic variation in Paramecium revealed by mitochondrial cytochrome C oxidase I sequences[J].Journal of Eukaryotic Microbiology,2006,53(1):20-25.
[47]Webster B L,Emery A M,Webster J P,et al.Genetic Diversity within Schistosoma haematobium:DNA Barcoding Reveals Two Distinct Groups[J].Plos Neglected Tropical Diseases,2012,6(10):10.