Cloning and sequencing of variable region gene of McAb H7 againsthantaan virus

  Objective   To clone and analyze the variable gene of monoclonal antibody H7 againsthantaan virus.

  Methods   According to amino acid consensus sequences of mouse immunoglobulins(Igs)as described by Kabat et al, degenerated primers were designed and synthesized.The nucleotide sequences were derived by using the codons preferentially used in murine Ig variable region(V)genes as listed in the Kabat database.Based on RT-PCR technology, succeeding in preparing mRNA、amplifying cDNA and in cloning Ig heavy(VH)and light(VK)chain variable domains from murine hybridoma H7, which secreted antibody specifically binding to HFRSV, and the variable genes were cloned into T7-sequencing vector respectively.

  Results   the subsequent sequence analyses were performed with a DNA-STAR sequence analysis program, and the nucleotide and predicted amino acid sequences indicated that H7-VK gene consisted of 357 bp and 119 amino acids (JK4), 360 bp and 120 amino acids of H7-VH gene(JH4)respectively.The sequences were submitted to the GenBank database and they were given the following accession numbers: AY245601 and AY245602.

  Conclusion   the results would be potentially useful in constructing engineering antibody(such as genetic evolvement and affinity maturation in vitro)as thera-peutic reagents for HFRS patients.