RESUMO
OBJECTIVE: To report on a novel KIR3DL3 allele identified in a southern Han Chinese individual. METHODS: Peripheral blood sample was collected from a voluntary blood donor with inconclusive result by KIR3DL3 sequence-based typing (SBT). Total mRNA was extracted and subjected to reverse transcription to obtain KIR3DL3 cDNA, which was then amplified by PCR with a pair of KIR3DL3-specific primers. The product was subjected to cDNA cloning and sequencing. RESULTS: cDNA cloning and sequencing have identified a wide-type KIR3DL3*00802 allele and a novel KIR3DL3*064 allele. The latter differed from KIR3DL3*00601 by a missense variant at codon 374[c.1184 C>T (p.Thr374Ile)] in exon 9. The novel KIR3DL3 allele has been officially assigned by the KIR subcommittee of World Health Organization Nomenclature Committee for factors of HLA system. CONCLUSION: cDNA cloning and sequencing may be used to distinguish inconclusive results in KIR3DL3 SBT in order to identify novel KIR alleles.
Assuntos
Alelos , Receptores KIR/genética , Sequência de Bases , Clonagem Molecular , Códon , DNA Complementar/genética , Humanos , Mutação de Sentido Incorreto , Análise de Sequência de DNARESUMO
OBJECTIVE: To list the key points for quality control during HLA-A, B, C, DRB1 and DQB1 allele typing by taking consideration of hardware, software and experimental procedures. METHODS: A total of 10 167 samples from randomly selected healthy blood donors and donor-recipient pairs from Shenzhen were typed for exons 2-4 of HLA-A, B, C, exon 2 of HLA-DRB1, and exons 2 and 3 of HLA-DQB1 by PCR- sequence-based typing. For 56 cases whose forward and reverse sequences were inconsistent, the samples were re-checked by a PCR-sequence specific oligonucleotide probe method. Novel alleles not included in the IMGT/HLA database were cloned and sequenced using in-house primers. RESULTS: Eight novel HLA alleles were identified. A table for key positions of single nucleotide polymorphisms (SNPs) were generated, which summarized the key points for quality control during HLA-A, B, C, DRB1 and DQB1 allele typing. Among the listed SNPs, 3 were located at the HLA-A locus, 8 were at the HLA-B locus, 6 were at the C locus, 6 were at the DQB1 locus, and 4 were at the DRB1 locus. To ensure the quality control, an unique sample number for DNA transferring tubes in the process of experiment should be considered. CONCLUSION: A protocol for quality control should be enforced by checking all of the key points. The SNPs and critical control points of the alleles should be examined to ensure the accuracy of HLA typing results.
Assuntos
Teste de Histocompatibilidade/métodos , Adulto , Alelos , Sequência de Bases , Primers do DNA/genética , Éxons , Feminino , Genótipo , Antígenos HLA-A/genética , Cadeias HLA-DRB1/genética , Humanos , Masculino , Dados de Sequência Molecular , Reação em Cadeia da Polimerase , Adulto JovemRESUMO
HLA-B*40:01:45 differs from HLA-B*40:01:02 by a single nucleotide change in exon 1, 33 G > A (codon -14 CTG > CTA).
Assuntos
Genes MHC Classe I , Hepatite B , Alelos , Sequência de Bases , Antígenos HLA-B/genética , Hepatite B/genética , Humanos , Análise de Sequência de DNARESUMO
BACKGROUND: There are no current estimates of the residual risks of transmission by blood of hepatitis B virus (HBV) or hepatitis C virus (HCV) and human immunodeficiency virus (HIV) in China. Such estimates are an essential prerequisite to monitoring and improving transfusion safety as well as supporting evidence based assessment of the value of implementing new screening interventions. STUDY DESIGN AND METHODS: Viral screening data for donors from Shenzhen, China, for the period 2001 to 2004, were retrospectively analyzed. The data were applied to a published model to estimate the residual risk of transmitting HIV, HBV, and HCV by blood transfusion in Shenzhen, as well as to assess the residual risk reduction value of various new tests. RESULTS: The point estimates for the combined 2003 and 2004 period calculate as 1 in 17,501 for HBV, 1 in 59,588 for HCV, and 1 in 903,498 for HIV. The predicted yield for improved hepatitis B surface antigen (HBsAg) assays, minipool (MP) nucleic acid testing (NAT), and individual-donation (ID) NAT was 6.9, 9.5, and 28.3 per million donations, respectively. The predicted yield for implementing a fourth-generation HCV (antigen-antibody) or MP NAT assay was 13.4 or 14.7 per million donations, respectively. For HIV, the predicted yield for implementing a fourth-generation HIV (antigen-antibody) or MP NAT assay was markedly smaller, 0.25 or 0.65 per million donations, respectively. CONCLUSIONS: Relative to that reported for Western blood systems, the prevalence and the residual risk of HBV and HCV are high, whereas HIV is comparable. Pending a formal cost-effectiveness study for NAT, implementing improved HBsAg and combination HCV antibody-antigen assays in Shenzhen would markedly reduce the residual risk.