PCR-based markers for the genotype identification of flavonoid- 3′,5′-hydroxylase genes governing floral anthocyanin biosynthesis in commercial petunias

Abstract

The major floral pigment of petunia, which is either a cyanidin-series (cyanidin and peonidin derivatives) or a delphinidin-series anthocyanin (delphinidin, petunidin and malvidin derivatives), is determined by the function of a flavonoid-3′,5′-hydroxylase that is encoded by two independent loci, Hf1 and Hf2. At the Hf2 locus, the recessive allele hf2-1 had a deletion of about 200 base pairs in the first intron and eight missense mutations by base substitution in the exons. The transcript level of hf2-1 was markedly lower than that of Hf2. We developed PCR-based markers to determine genotypes of the Hf1 and Hf2 loci. This technique allows amplified fragments from the Hf1, hf1-2 and hf1-3 alleles, as well as from the Hf2 and hf2-1 alleles, to be distinguished from one another. These PCR-based analyses showed no discrepancy between the Hf1/Hf2 loci genotypes and the major floral anthocyanidin phenotypes in the 129 commercial petunias we examined. Another reported recessive mutant allele at the Hf1 locus, caused by the insertion of an Spm-like transposable element in the second exon (hf1-1), was not found in the commercial petunias.