Chinese Journal of Tissue Engineering Research ›› 2011, Vol. 15 ›› Issue (5): 878-881.doi: 10.3969/j.issn.1673-8225.2011.05.028

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Identification of a human new allele human leucocyte antigen-B*15:133 and its prokaryotic expression

Wang Lin1, Song Chang-xing2, Zhang Zhi-xin2   

  1. 1Beijing Tuberculosis and Thoracic Tumor Research Institute, Beijing  101149, China
    2Beijing Red Cross Blood Center, Beijing  100088, China
  • Received:2010-07-07 Revised:2010-08-23 Online:2011-01-29 Published:2011-01-29
  • Contact: Zhang Zhi-xin, Professor, Beijing Red Cross Blood Center, Beijing 100088, China
  • About author:Wang Lin☆, Doctor, Technician, Beijing Tuberculosis and Thoracic Tumor Research Institute, Beijing 101149, China cod_wang@hotmail. com

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Abstract:

BACKGROUND: In total 4 633 alleles have been reported in world up to April 2010. More than 200 new alleles have been found in China, most of which need to be further studied.
OBJECTIVE: Serologic and genetic studies were performed to confirm the identity of a new human leucocyte antigen (HLA) allele within the Chinese population to construct an expression vector containing the heavy chain of HLA-B*15:133 in a prokaryotic system and to identify its activity.
METHODS: During routine typing work, genomic DNA was typed by PCR-SSO and PCR-SBT. An ambiguous result was found and identified as a novel allele by Serologic and DNA sequence analysis. The extra-membrane gene fragment of HLA-B*15:133 (digested with BamHⅠand HindⅢ) was inserted into vector pET 30a(+). The expression vector pET30a(+)-B*15:133 was transfected into the BL21(DE3) cells, and western-blotting was used to identify its expression.
RESULTS AND CONCLUSION: A new allele which had one nucleotide substitution at position 419 from C to T (condon116 TCC->TTC) was identified, resulting in an amino acid change from Serine (S) to Phenylalanine (F). Serologic specificity is B71 (70) genetic and serological analysis indicated that the novel HLA-B allele of the donor was inherited from his mother. The restriction endonuclease digestions and PCR suggested the expression vector pET 30a(+)-B*15:133 was constructed successfully. A Western-blot identified the extra-membrane polypeptide chain of HLA-B*15:133. The expression vector pET30a(+)-B*15:133 can express the extra-membrane polypeptide chain of HLA-B*15:133 in BL21(DE3).

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