豚鼠MCHR2基因外显子1的克隆及序列分析

目的 探明豚鼠体内MCHR2基因的表达情况, 为构建动物模型提供实验依据.方法 利用逆转录-多聚酶链反应(RT-PCR)方法,从新鲜豚鼠大脑组织的总RNA中扩增MCHR2基因的全长cDNA编码区序列,从豚鼠脑组织中提取基因组DNA,PCR扩增MCHR2基因外显子1的序列,克隆入pUCm-T载体后进行序列分析.结果 RT-PCR法未获得目的 片段;由基因组DNA扩增出的MCHR2基因外显子1的序列片段长183 bp,与GenBank登录的人、猴、白鼬、犬的MCHR2序列比对,在第47位后面多了一个碱基--A,由于移码错译而导致提前出现终止密码,从而缩短了预测的开放阅读框架.结论 豚鼠体内的MCHR2基因是无功能的假基因.

Cloning and sequence analysis of guinea pig MCHR2 gene exon 1

Objective To study the expression of MCHR2 gene in guinea pig and to provide evidence for animal model construction. Methods The full-length MCHR2 cDNA fragment was amplified from the total RNA of fresh guinea pig brain tissue by RT-PCR. Then the MCHR2 gene exon 1 was amplified through PCR from the genomic DNA of guinea pig brain tissue and cloned into pUCm-T vector and sequenced. Results The full-length MCHR2 cDNA fragment was not obtained by RT-PCR. The DNA fragment of MCHR2 gene exon 1 was 183 bp in length. As compared with MCHR2 sequence of human, rhesus monkey, ferret and dog in GenBank, a frameshift in guinea pig was introduced (additional A after the 47th bp), which placed a premature termination codon shortly after the frameshift, shortening the predicted open reading frame. Conclusion Guinea pig contains nucleotide sequence alteration that renders MCHR2 non-functional.