小鼠脂肪组织脂联素基因编码区的克隆及序列分析

Cloning and Sequencing of Mouse ACRP30 Coding Sequence from Adipose Tissue

目的:克隆小鼠脂肪组织脂联素(ACRP30)编码区基因并进行序列分析。方法:用TRIzol试剂从小鼠脂肪组织提取总RNA,经RT-PCR方法扩增全长的ACRP30cDNA片段,将PCR产物克隆于pGEM-T载体,对重组质粒pGEM-ACRP30进行序列验证。结果:小鼠脂肪组织ACRP30基因编码序列不同于来自3T3-L1脂肪细胞的编码序列,IRM-2(InstituteofRadiationMedicine-2)小鼠和C57BL/6J小鼠脂肪组织的ACRP30基因编码序列相同。337位点上的碱基A被G置换导致在113位的蛋氨酸被缬氨酸取代。来自脂肪组织的ACRP30基因编码序列已被提交到GenBankTM数据库,C57BL/6J小鼠的收录编号为AY749429,IRM-2小鼠为AY754346。结论:小鼠脂肪组织ACRP30编码序列是ACRP30序列的新成员,cDNA克隆的构建为进一步的蛋白表达和生物学活性研究奠定了基础。

Objective： Cloning for coding sequence of ACRF30 from mouse adipose tissue and analysis of its sequence. Methods： Total RNA was isolated from mouse adipose tissue using TRIzol reagent. The full-length ACRP30 cDNA fragment was amplified by reverse transcription-PCR （RT-PCR）. The PCR product was cloned into pGEM-T vector to confirm the sequence of recombination plasmid pGEM-ACRP30. Results： The coding sequence of ACRP30 derived from mouse adipose tissue was different from the coding sequence derived from 3T3-LI adipocytes. The coding sequences of ACRP30 derived from mouse adipose tissue of Institute of Radiation Medicine-2 （IRM-2） mouse and C57BL/6J mouse are identical, and A substituted by G at nucleotide 337 leaded to amino acid substitution from methionine to valine at position 113. The coding sequence of ACRP30 derived from adipose tissue had been submitted to GenBankTM, the Accession Numbers are AY749429 for C57BL/6J mouse and AY754346 for IRM-2 mouse. Conclusion： The coding sequence ACRP30 derived from mouse adipose tissue is a new member of ACRP30. The construction of cDNA clone provides a basis for further research on protein expression and biological activities.