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目的:分离和鉴定大鼠巨噬细胞环加氧酶-2剪接异构体MRNA片段,并分析其可能意义。方法: 采用RT-PCR和测序分析从大鼠巨噬细胞扩增、分离环加氧酶-2剪接异构体CDNA片段,并推导环加氧酶-2剪接异构体氨基酸序列,利用生物信息软件对氨基酸序列进行分析。结果:大鼠肺和血管平滑肌组织及腹腔巨噬细胞均发现COX-2目的条带(253 BP),但腹腔巨噬细胞则出现了一条新的电泳带(422 BP),经克隆和测序证实,该条带除目的条带的COX-2 DNA的第7、8外显子序列外,还包含第7、8外显子间的内含子序列,即保留的内含子(169 BP)。根据阅读框推测,在保留的内含子第21-23碱基出现终止密码。结论:首次发现大鼠腹腔巨噬细胞中存在 COX-2基因的选择性剪接异构体(GENBANK ACCESSION NUMBER:BU500100),其生物学意义有待进一步研究。
OBJECTIVE: To isolate and identify the MRNA fragment of the cyclooxygenase-2 splicing isoform of rat macrophage and analyze its possible significance. METHODS: The cDNA of CDNA fragment of cyclooxygenase-2 splice variant was amplified from rat macrophages by RT-PCR and sequencing analysis. The amino acid sequence of the cyclooxygenase-2 splice variant was deduced. The bioinformatics The software analyzes the amino acid sequence. RESULTS: The target band (253 BP) of COX-2 was found in rat lung and vascular smooth muscle and in peritoneal macrophages. However, a new electrophoresis band (422 BP) appeared in peritoneal macrophages, which was confirmed by cloning and sequencing , In addition to the seventh and eighth exon sequences of the COX-2 DNA of the target band, contains intron sequences between exons 7 and 8, ie, the intron retained (169 BP) . Based on the reading frame speculation, a stop codon was found in the conserved intron 21-23 bases. CONCLUSION: For the first time, it is found that COX-2 alternative splicing isoforms exist in rat peritoneal macrophages (GENBANK ACCESSION NUMBER: BU500100), and its biological significance remains to be further studied.