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1、南京农业大学学报 2004,27(1):15～19JournalofNanjingAgriculturalUniversity三角叶滨藜甜菜碱醛脱氢酶(BADH)基因的克隆及序列分析1,2132423何晓兰,侯喜林,吴纪中,刘桂华,钦佩,朱卫民(11南京农业大学作物遗传与种质创新国家重点实验室,江苏南京210095;21江苏省农业科学院遗传生理研究所,31粮食作物研究所,江苏南京210014;41南京大学生物技术研究所,江苏南京210093)摘要:以藜科植物山菠菜甜菜碱醛脱氢酶(BADH)基因的保守区设计引物,通过RT2PCR技术,从三角叶滨藜cDNA中分离了两
2、个BADH基因片段BADH3和BADH5,并利用巢式PCR、cRACE以及3′2RACE获得全长BADH3。测序结果表明,BADH3和BADH5之间的核苷酸同源性为81%,其推测的氨基酸序列同源性80%。全长BADH3核苷酸序列长1815bp,79～1578bp为一个开放阅读框架,推测的氨基酸序列全长500个氨基酸残基,相对分子质量为54700,pI为515。其核苷酸序列和推测的氨基酸序列与山菠菜同源性分别为95%和93%。由三角叶滨藜3′端部分BADH基因的克隆gBADH1和gBADH2推测的外显子序列分别与BADH5和BADH3完全一致,且gBADH1和gB
3、ADH2的内含子插入位置和大小有较大差异。表明三角叶滨藜BADH基因是一个多基因家族,至少存在2个成员。关键词:三角叶滨藜;甜菜碱 脱氢酶;基因;克隆;聚合酶链式反应中图分类号:Q949174511;Q781文献标识码:A文章编号:10002030(2004)01001505Molecularcloningandsequenceanalysisofbetaine2aldehydedehydrogenase(BADH)inAtriplextriangularis1,2132423HEXiao2lan,HOUXi2lin,WUJi2zhong,LIUGui2hua,
4、QINPei,ZHUWei2min(11NationalKeyLaboratoryofGeneticsandGermplasmEnhancement,NanjingAgriculturalUniversity,Nanjing210095,China;21InstituteofAgrobiologicalGeneticsandPhysiology;31InstituteofFoodandCrop,JiangsuAcademyofAgriculturalSciences,Nanjing210014,China;41InstituteofBiotechnology,N
5、anjingUniversity,Nanjing210093,China)Abstract:BADH3andBADH5,twopartialBADHcDNAswereclonedfromAtriplextriangularisthroughRT2PCRtechniquebyspecificprimersdesignedonthebasisofBADHgeneonAtriplexhortensis.ComparisonbetweenBADH3andBADH5showed81%identityfornu2cleotidesequencesand80%identity
6、fordeducedaminoacidsequences,respectively.Moreover,a1815bpfull2lengthcDNAsequenceofBADH3wasobtainedusingnestedPCR,cRACEand3′2RACEtechnique.FurtheranalysesonBADH3indicatedthatanopenreadingframe(ORF)of1500bpcontainedinthisgeneandpredictedtoencodeaproteinofMr54700with500aminoacids.Thenu
7、cleotideandthededucedaminoacidsequencesofthisfull2lengthBADH3show95%and93%identitytoAtriplexhortensisBADHgene,respectively.Genomicclones(gBADH1andgBADH2)of3′endofBADHgenewereamplifiedandisolatedfromAtriplextriangularis,whichintronsshowsignificantdifferenceinthelengthandposition.Thede
8、ducedtranscr
