Restriction digests of the clone give the following sizes (kb): BamHI/HindIII--3.25, 0.6; BamHI--3.8; PstI--2.8, 0.71; HindIII--3.8; EcoRI--3.8. The insert covers the entire alpha chain coding region and leader. There are typically two copies of the gene per haploid genome. HindIII detects a 1.7 kb length polymorphism which is also detected by BamHI with fragment sizes of 7.5 kb and 9.2 kb. The HindIII polymorphism is also detected by AvaII with fragment sizes of 4.4 kb (freq = 0.65) and 2.7 (freq = 0.35). The AvaII RFLP also appears to be in linkage equilibrium with a TaqI RFLP at the CETP locus. There is a polymorphic EcoRI site about 4 kb 3' to Hp2 which reduces the allele size to 10.1 (indistinguishable from the Hp1 allele). For the XbaI polymorphism, the 5.8 kb constant band is from the HPR sequences, the 4.6 kb band is from the Hp2 or Hp1F alleles, and the 4.0 kb band is from the Hp1S allele. For the HindIII polymorphism, the 3.4 kb constant band is from the HPR sequences, the 4.9 kb band is from the Hp2 allele, and the 3.2 kb band is from the Hp1S or Hp1F alleles. For the EcoRI polymorphism, the 8.3 kb constant band is from the HPR sequences, the 11.8 kb band is from the Hp2 allele, and the 10.1 kb band is from the Hp1S or Hp1F alleles. A small HindIII/PstI linker was used in the process of inserting the PstI/BamHI fragment into the HindIII and BamHI sites of pAT153. |
