Abstract:

Low temperature and cold damage are important factors limiting the regional distribution and yield of cultivated rice， and this seriously affects food security in China. The C-repeat binding factor cold-regulated gene （CBF-COR） pathway with CBF/DREB1 transcription factors as the core component is an important signal transduction pathway involved in the cold acclimation of rice. A total of 71 members of the CBF/DREB1 gene subfamily were identified based on whole genome sequencing results and hidden markov model （HMM） searches of nine species in the rice （Oryza） genus. All genes were intron-free and consisted of a single exon. Most of them encoded weakly acidic proteins. All putative DREB1 proteins had a grand average of hydropathicity （GRAVY） of <0. A phylogenetic analysis divided into three groups： Group I （DREB1A/1B/1H）， Group Ⅱ（DREB1C/1E/1F/1G）， and Group Ⅲ （DREB1D/1I/1J）. The motif composition differed among the groups， although all of the putative proteins contained the characteristic AP2 domain and flanking sequence. An adaptive evolutionary analysis of japonica subspecies and other rice orthologuous gene pairs showed that the nonsynonymous nucleotide substitution rate/synonymous nucleotide substitution rate （Ka/Ks） values for the gene pairs OsDREB1A/ObDREB1A， OsDREB1D/OnDREB1D，OsDREB1I/OsIDREB1I， and OsDREB1J/OsIDREB1J were greater than 1， indicating that they were under positive selection pressure. The types and numbers of promoter elements were similar in orthologs， but differed between paralogs. Real-time quantitative PCR analyses were conducted for 7-day-old seedlings of Nipponbare， 93-11 and Dongxiang wild rice after 2 hours of cold treatment at 4 ℃. The results showed that， during the early cold response， some DREB genes were activated （DREB1A/1B/1C/1E/1F/1G/1H） and others were inhibited （DREB1D/1I/1J）. Among the cold-activated genes in Nipponbare， 93-11， and Dongxiang wild rice， DREB1B/1G/1H were the first and the fastest to respond to cold. All of these three genes had at least one copy of the CAMTA transcription factor binding element CM2 （CCGCGT） in their promoter regions. The transcript levels of OrDREB1B/1C/1E/1F/1G/1H in Dongxiang wild ricewere higher than those of their homologs in cultivated rice after 4 hours of cold treatment at 4℃. The results of this molecular evolutionary study on the CBF/DREB1 gene subfamily in nine species in the Oryza genus provide a preliminary basis for the mining and utilization of low-temperature resistance genes.

Key words: wild rice, CBF/DREB1, motif, selective pressure, promoter analysis