齿瓣石斛的位点特异性PCR鉴别

目的设计出齿瓣石斛的位点特异性鉴别引物,仅通过PCR就能完成对齿瓣石斛真伪进行准确鉴别。方法根据齿瓣石斛及其他枫斗类、黄草类石斛的rDNA ITS序列数据库,设计了齿瓣石斛的位点特异性PCR鉴别引物JB-Chiban-01S和JB-Chiban-01X。然后,对38种石斛属植物模板DNA进行了PCR扩增,阳性者即为齿瓣石斛正品。结果当退火温度设定为58℃时,只有齿瓣石斛的模板DNA能被扩增出来,而其他的37种石斛属植物均为阴性。该鉴别反应重复性好,已在鉴别齿瓣石斛时发挥重要作用。结论运用位点特异性鉴别引物能成功地对齿瓣石斛进行PCR鉴别,与DNA测序鉴别方法相比,位点特异性PCR具有高效、准确、简便、省时等优点。

ALLELE-SPECIFIC DIAGNOSTIC PCR AUTHENTICATION OF D.DEVONIANUM FROM OTHER DENDROBIUM SPECIES

AIMTo design a pair of allele-specific diagnostic primers for authenticating D.devonianum from other Dendrobium species by PCR. METHODSBased on rDNA ITS sequences database of D.devonianum and other Dendrobium species of Fengdous and Huangcaos, an allele-specific diagnostic primer pair, JB-Chiban-01S and JB-Chiban-01X, was designed. Diagnostic PCRs were performed using the primer pair with the total DNAs of the original plants of 38 Dendrobium species as templates. RESULTSWhen the annealing temperature was raised to 58℃, only the template DNA of D.devonianum could be amplified whereas the diagnostic PCR of the rest 37 species were all negative. The diagnostic PCR have been repeated many times and played an important role in authenticating D.devonianum in China. CONCLUSIONThe diagnostic PCRs can authenticate D.devonianum from other Dendrobium species efficiently. Compared with the anthentication method by sequencing DNA fragments, the allele-specific diagnostic PCR was not only simple and timesaving but also practical and effective.