Isolation of circadian-associated genes in Brassica rapa by comparative genomics with Arabidopsis thaliana.
Elucidation of the roles of circadian associated factors requires a better understanding of the molecular mechanisms of circadian rhythms, control of flowering time through photoperiodic pathways, and photosensory signal transduction. In Arabidopsis, the APRR1 quintet, APRRs 1, 3, 5, 7, and 9, are known as central oscillator genes. Other plants may share the molecular mechanism underlying the circadian rhythm. To identify and characterize these circadian response genes in Brassica crops whose genome was triplicated after divergence from Arabidopsis, we identified B. rapa BAC clones containing these genes by BLAST analysis of B. rapa BAC end sequences against the five corresponding Arabidopsis regions. Subsequent fingerprinting, Southern hybridization, and PCR allowed identification of five BAC clones, one for each of the five circadian-related genes. By draft shotgun sequencing of the BAC clones, we identified the complete gene sequences and cloned the five expressed B. rapa circadian-associated gene members, BrPRRs 1, 3, 5, 7, and 9. Phylogenetic analysis revealed that each BrPRR was orthologous to the corresponding APRR at the sequence level. Northern hybridization revealed that the five genes were transcribed at distinct points in the 24 hour period, and Southern hybridization revealed that they are present in 2, 1, 2, 2, and 1 copies, respectively in the B. rapa genome, which was triplicated and then diploidized during the last 15 million years.
Kim
Jin A
JA
Brassica Genomics Team, National Institute of Agricultural Biotechnology (NIAB), Suwon 441-707, Korea.
Yang
Tae-Jin
TJ
Kim
Jung Sun
JS
Park
Jee Young
JY
Kwon
Soo-Jin
SJ
Lim
Myung-Ho
MH
Jin
Mina
M
Lee
Sang Choon
SC
Lee
Soo In
SI
Choi
Beom-Soon
BS
Um
Sang-Hee
SH
Kim
Ho-Il
HI
Chun
Changhoo
C
Park
Beom-Seok
BS
D000595:Amino Acid Sequence; D017360:Arabidopsis / Q000235:genetics* / Q000378:metabolism; D029741:Brassica rapa / Q000235:genetics* / Q000378:metabolism; D002940:Circadian Rhythm / Q000502:physiology*; D018745:Genome, Plant; D023281:Genomics; D008969:Molecular Sequence Data; D010802:Phylogeny