England Biolabs, Ipswich, MA, USA), in a 20 μl reaction assay containing 1X NEBuffer 1 and 1X BSA at 37 °C for 1 h. Digested and non-digested PCR products were loaded on 3% agarose gel in 1X TBE. The short ‘s’ and the long ‘l’ alleles (as well as the Stin2.9, Stin2.10 and Stin2.12 alleles not used in this study) were determined from the non-digested PCR products with fragments of 469 bp, 512 bp, 250 bp, 267 bp and 300 bp respectively. The rs25531 A/G polymorphism was determined after HpaII digestion according to the following fragment lengths (LA: 512 bp, LG: 402 + 110 bp, SA: 469 bp and SG: 402 + 67 bp). The more common LA allele is associated with the reported higher basal activity, whereas the less common LG allele has transcriptional activity no greater than the S. Therefore, it is suggested that in tests of association the LG alleles should be analyzed along with the S alleles.
