人BAFF cDNA的克隆及原核表达载体的构建

目的 克隆人BAFF（B cell activating factor belonging to the TNF family)全长及128－285片段的cDNA并构建其原核表达载体，为表达和检测其生物学活性做准备。方法 提取HL－60细胞总RNA，经RT－PCR扩增编码人BAFF全长及128－285氨基酸残基的cDNA序列，并将其克隆至载体pUC18和pUC19进行序列测定，然后构建于新型原核表达载体pQE-80L中。结果 RT－PCR扩增得到了858bp和477bp的DNA片段，序列分析与GenBank中报道的编码BAFF全长及128－285氨基酸残基的cDNA序列完全一致。结论 BAFF cDNA的克隆及其原核表达载体的构建，为进一步表达和检测其生物学活性奠定了基础。

Cloning of human BAFF cDNAs and the construction of their prokaryotic expression vector

Objective To clone human BAFF cDNAs and construct their prokaryotic expression vector for expressing and examining their activity. Methods The total RNA was extracted from HL 60 and cDNAs were amplified to encode human BAFF full length and 128-285 amino acid resides by using RT PCR and ligated into vectors pUC18 and pUC19 for sequence analysis. Then the identified cDNAs were ligated into the new type of prokaryotic expression vector pQE 80L. Results The acquired DNA fragments by RT PCR amplification 858 bp and 477 bp cDNA were completely the same as the sequence reported in GenBank. Conclusion Cloning of BAFF cDNA and the construction of its prokaryotic expression vector have provided fundamental proof for expressing BAFF and examining its biological activity.