基于COI与SRY序列建立梅花鹿、马鹿及其杂交鹿茸的分子鉴别方法

Identification of Cervus nippon,C. elaphus and their hybridize samples based on COI and SRY gene

为了建立梅花鹿、马鹿及其杂交鹿茸的特异性PCR鉴别方法。采集不同来源的梅花鹿、马鹿鹿及其杂交鹿的鹿角或鹿茸样品共48份,其中24份为市场流通的待测样品。分别利用COI与SRY基因序列作为其父母本的鉴别标记,对已知样品的COI与SRY基因进行扩增、测序,进行同源比对后根据其变异位点设计特异鉴别引物,分别基于COI与SRY基因建立对梅花鹿、马鹿及其杂交鹿的鹿茸的特异PCR鉴定方法,并随机对市场流通的24份待测鹿茸样品进行鉴别分析。该实验所构建双位点特异PCR体系,基于COI的鉴别位点,可通过PCR扩增获得232 bp的梅花鹿特异片段,而产生518 bp的马鹿特异片段;而基于SRY的鉴别位点,通过PCR扩增获得803 bp的梅花鹿特异片段,而产生425 bp的马鹿特异片段。随机抽取市场流通中的24个待测鹿茸样品,经鉴定有3个样品属于马·花杂交的鹿茸样品。该实验建立对梅花鹿、马鹿及其杂交鹿的鹿茸的PCR鉴别方法简便、可靠。

For rapid identification of Ce^us nippon, C. elaphus and their hybridize samples, the specific PCR for mutual autilcnti- cation of them was established based on the SNPs in CO1 and SRY sequence. C. nippon, C. elaphus and their hybridize samples were collected from different origins, total DNA of 24 identified samples were extracted, and the CO1 and SRY gene was seqenced. SNPs in the CO1 and SRY sequences of the samples were found by Clustul X 2. 1 program. Primers for identifying C. nippon and C. elaphus were designed according to the SNP site, two muhi-PCR reaction system were established to identify them. In addition, 2d samples which were randomly collected in different herbal medicine market were identified. The band special for C. nippon （232 bp）and band special for C. elaphus （518 bp） based on CO1 sequence,and the band special for C. nippon （803 bp）and band special for C. elaphus （425 bp） based on SRY sequence, were found using multi-PCR reaction, and three of the twenty-four samples were identified as the hybridize samples. The muhi-PCR reaction system could be used to identify C. nippon, C. elaphus and their hybridize samples.