EVIDENCE THAT THE PHYTOCHROME GENE FAMILY IN BLACK COTTONWOOD HAS ONEPHYA LOCUS AND 2 PHYB LOCI BUT LACKS MEMBERS OF THE PHYC F AND PHYE SUBFAMILIES/

The phytochrome photoreceptors play important roles in the photoperiod ic control of vegetative bud set, growth cessation, dormancy induction , and cold-hardiness in trees. Interestingly, ecotypic differences in photoperiodic responses are observed in many temperate-zone tree speci es. Northern and southern ecotypes of black cottonwood (Populus tricho carpa Torr. & Gray), for example, exhibit marked differences in the ti ming of short-day-induced bud set and growth cessation, and these resp onses are controlled by phytochrome. Therefore, as a first step toward determining the molecular genetic basis of photoperiodic ecotypes in trees, we characterized the phytochrome gene (PHY) family in black cot tonwood. We recovered fragments of one PHYA and two PHYB using PCR-bas ed cloning and by screening a genomic library. Results from Southern a nalyses confirmed that black cottonwood has one PHYA locus and two PHY B loci, which we arbitrarily designated PHYB1 and PHYB2. Phylogenetic analyses which included PHY from black cottonwood, Arabidopsis thalian a and tomato (Solanum lycopersicum) suggest that the PHYB/D duplicatio ns in these species occurred independently. When Southern blots were p robed with PHYC, PHYE, and PHYF heterologous probes, the strongest ban ds that we detected were those of black cottonwood PHYA and/or PHYB. T hese results suggest that black cottonwood lacks members of the PHYC/F and PHYE subfamilies. Although black cottonwood could contain additio nal PHY that are distantly related to known angiosperm PHY, our result s imply that the PHY family of black cottonwood is less complex than t hat of other well-characterized dicot species such as Arabidopsis and tomato. Based on Southern analyses of five black cottonwood genotypes representing three photoperiodic ecotypes, substantial polymorphism wa s detected for at least one of the PHYB loci but not for the PHYA locu s. The novel character of the PHY family in black cottonwood, as well as the differences in polymorphism we observed between the PHYA and PH YB subfamilies, indicates that a number of fundamental macro-and micro evolutionary questions remain to be answered about the PHY family in d icots.