Identification of JK gene mutations in Jknull variants.
(A) Left, schematic representation of the Jk(Δ6) DNA genotyping assay by allele-specific PCR using primers SP-4 and AS-5. The invariant G nucleotide at the acceptor splice-site of intron 5 in JK*Aor JK*B alleles is replaced by an A in the Jk(Δ6)null allele. Right, the 221-bp PCR product was amplified from the Jk(Δ6) mutant but not from the wt Jk, Jk(a+b−) or Jk(a−b+), Jk(Δ7), and Jk(S291P) samples demonstrating the specificity of the assay. (B) Left, schematic representation of the Jk(Δ7) DNA genotyping assay by PCR-RFLP using primers SP-6, SP-7, and AS-8. The invariant G nucleotide at the donor splice-site of intron 7 inJK*A or JK*B alleles is replaced by a T in theJk(Δ7) null allele, which creates a Mse I restriction site. Right, the 76-bp PCR product encompassing the splice-site mutation was cleaved by Mse I into 48- and 28-bp fragments in the Jk(Δ7) mutant. As expected the PCR products from wt Jk, Jk(a+b−) or Jk(a−b+), Jk(Δ6), and Jk(S291P) samples were uncut. Single base substitution and intronic sequence are shown by a star and in lower case, respectively. Size of fragments (bp) are given on the right.
