摘要
本研究用 1 6S 2 3SrDNA间的ITS序列通用引物L1 ( 5′ AGTCGTAACAAGGTAGCCGT 3′)和L2 ( 5′ GTGCCAAGGCATCCACC 3)扩增甜菜银叶病菌 (Curtobacteriumflaccumfacienspv .betae,Cfb)和其它相近细菌的基因组DNA ;并对其PCR产物进行回收、克隆和测序 ,将所获序列和其它已报道的细菌内源转录间隔区 (InternallyTranscribedSpacer,ITS)序列进行多重比较后设计出Cfb的特异性引物B1 ( 5′ GGCCTCGTGTTGTCCCTTATC 3′)和B2 ( 5′ GTCACCAATCAA CAACCCGAG 3′)。此引物可以从Cfb中扩增出 387bp的特异性片段 ,而其余参试的 2 1个细菌PCR反应结果均为阴性。该方法可以应用于病害防治工作中的Cfb快速。
A PCR protocol was developed that specifically detected Curtobacterium flaccumfaciens pv. betae (Cfb), the causatine agent of silvering disease of red beet. Generic PCR products from the internally transcribed spacer (ITS) region of 16S～23S ribosomal DNA of Cfb and other related bacteria were cloned and sequenced. Based on a multiple sequences alignment among these obtained sequences and other nonredundant highly homologous sequences from database, two Cfb specific PCR primers were designed, B1(5′ GGCCTCGTGTTGTCCCTTATC 3′)/B2 (5′ GTCACCAATCAACAACCCGAG 3′). These two oligonucleotides primed the specific amplification of a 387bp DNA product from genomic DNA sample of Cfb strain. Amplification was not observed with other 21 tested bacteria, including 5 strains of other Cur. f. pvs., 3 strains of Clavibacter species, 8 type strains of other different genus and 5 strains of saprophytic bacteria. The PCR protocol provides a rapid, reliable and economical tool for routine detection and identification of Cfb.
出处
《微生物学报》
CAS
CSCD
北大核心
2003年第2期271-275,共5页
Acta Microbiologica Sinica
基金
国家自然科学基金 (3 9970 481 )
中国农业科学院植物保护研究所病虫害生物学国家重点实验室 2 0 0 1年度开放基金 (pd3 )资助项目~~
关键词
甜菜银叶病菌
ITS分析
PCR检测
Curtobacterium flaccumfacies pv. betae, ITS analysis, PCR detection
