Characterization and expression of FLORICAULA/LEAFY homologues in Buddleja davidii

Buddleja, a cosmopolitan taxon of roughly 100 species, provides a unique model for studying inflorescence development at the molecular level. Great diversity in inflorescence architecture exists among Buddleja species, and numerous hybrids exist between and among these taxa. Breeding goals have included enhancement of floral architecture through increased panicle branching and total flowers per inflorescence. The B. davidii inflorescence is an indeterminate panicle of racemes, and several clones exhibiting enhanced inflorescence branching are known.;Homologues of the floral meristem identity genes FLORICAULA from Antirrhinum majus and LEAFY from Arabidopsis thaliana have been isolated from B. davidii in an effort to facilitate our understanding of the molecular contribution to inflorescence branching. Five full-length cDNAs were identified as FLO/LFY homologues. Nucleotide sequence identity among the five clones was at least 96%. Three clones shared 100% identity at both the nucleotide and deduced amino acid sequence level with the exception of gaped regions. These three appear to represent alternative splice forms of a single allele (BdFL1alpha, BdFL1beta and BdFL1gamma) and two others (BdFL2 and BdFL3) represent separate alleles. Nucleotide sequence homology of BdFL clones was 86% to 88% with FLO and 62% to 68% with LFY.;Qualitative expression of all five clones was examined in seven tissue types. Total RNA from meristems of shoots bearing 1--3, 4--6 or 7--9 nodes, terminal or lateral inflorescences, whole flowers or vegetative leaves was used to generate first-strand cDNA, and a PCR was carried out using the shared primers BdX1 and BdX2. Amplicons of all five clones together would resolve as only two bands following gel electrophoresis because of the identical or nearly identical number of base pairs in the amplified region. However, three out of the five clones possess a single NgoMIV restriction site 105 bp downstream of the BdX1 priming site. Therefore, to resolve individual fragments, each RT-PCR product was restricted with NgoMIV. Restriction products were analyzed on an ethidium bromide stained 10% polyacrylamide gel. Four of the five clones were detectable in five out of seven samples; only BdFL1gamma was not found in any sample at detectable levels.