Key result
PCR-RFLP using specific primers and endonucleases can unambiguously resolve all 22 DQB1 alleles and most of their possible heterozygotes, providing a reliable method for DQB1 typing.
PCR-RFLP is a simple, inexpensive, and reliable method for comprehensive DQB1 allele typing.
May support affordable DQB1 typing in select labs; leaves open its role versus sequencing in current practice.
The protocols represented in this report can resolve all 22 DQB1 alleles. The second exon of DQB1 was subjected to PCR using two group-specific primers to obtain DQB1 group 1 (DQ5 and DQ6) and group 2 (DQ2, DQ3, DQ4) specific amplified products, respectively. Three endonucleases, ApaI, BssHII and NciI, can provide typing of DQ5 and DQ6 based on easy-to-read uncleaved, cleaved and a combination of uncleaved/cleaved patterns. Similarly, two endonucleases, FokI and BgII can define the specificities DQ2, DQ3 and DQ4. Moreover, all 13 group 1 DQB1 alleles and all but one of their 78 possible heterozygotes can be unambiguously resolved using an extended panel of 10 endonucleases. The remaining pair of heterozygotes, DQB1*05031/0603 and 05032/0608, can however be resolved by double digestion with BsmFI and SfaNI. RsaI splits the previously unresolved alleles DQB1*0602 and 0603 in the amplified products of the modified primer SDQ-01. Fnu4HI can resolve DQB1*0606 from 0605. DQB1*0603, 0607 and 0608 can be resolved by SfaNI and the new endonuclease BsmFI. The comprehensive typing of group 2 DQB1 alleles can be achieved using five endonucleases. All 9 group 2 DQB1 alleles and all but one pair (DQB1*0301/0302 from DQB1*03032/0304) of 36 possible heterozygotes can be resolved. Thus, PCR-RFLP remains a simple, inexpensive and reliable method for DQB1 typing. The PCR-RFLP can be used for comprehensive DQB1 typing either independently or to complement the PCR-SSP and PCR-SSOP methods.
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Sengar et al. (1994) studied DQB1 allele typing. PCR-RFLP was evaluated on Resolution of DQB1 alleles. PCR-RFLP using specific primers and endonucleases can unambiguously resolve all 22 DQB1 alleles and most of their possible heterozygotes, providing a reliable method for DQB1 typing.
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