Expressing Characterization Of The Nine ABI5 Subfamily Members In Arabidopsis Thaliana

The ABI5 subfamily is a group of basic leucine zipper transcription factors that funtion in the maturation of late embry and respond to ABA and stress enviroments such as, drought, high salt, etc. There are nine members of the subfamily identified in Arabidopsis thaliana. Expressing tissues and period of the nine members and their roles in detail are not exactly understanded during plant growth and development. Moreover, molecular mechanism of their expressing regulations are unclear. Although some studies on the temporal and spatial expression patterns of the ABI5 subfamily members have been reported, there is no confirmedly conclusion. Therefore, it is still necessary that the expression patterns of the nine members and the involved signal pathways are more investigated. These researches are not only better to understand the functions of the ABI5 Subfamily genes, but much important to improve drought/salt-resistance of crops.The main results obtained in this paper are as follows:(1) The upstream sequences of the ABI5 subfamily members was analysised by an online software, PLACE, according to the nine member locations on chromosomes of Arabidopsis thaliana. Many cis-acting elements in their upstream sequences such as ARR1 AT, CARGCW8 GAT, GATABOX, GT1 CONSENSUS, MYB/MYC, AGL15(except for ABF3) were found as well as TATA-Box. These cis-acting elements would remind that the ABI5 subfamily members may regulated by ARR1, GT1, GATA, MYB/MYC and AGL15 transcription factors. PCR primers were designed based on Primer 5. The upstream sequences of the nine members were amplify and seven of them(ABF1, ABF2, ABF3, ABF4, DPBF2, DPBF3, DPBF4) were cloned into plant genetic transformation vectors, p CAMBIA1301 or p BIB-BASTA-GUS-GWR.(2) Histochemical staining analysis of the ProABF1::GUS, ProABI5::GUS and ProABF4::GUS transgenetic seedlings: Arabidopsis thaliana were transformed by the flower-dip method with Agrobacteria GV3101 containing recombinant plasmids of which the GUS gene was driven by the ABI5 subfamily member promoters. 8 ProABF1::GUS, 23 ProABI5::GUS and 2 ProABF4::GUS transgenetic plants were obtained. After GUS-staining, roots, cotyledons and leaves of ProABF1::GUS transgenic plants presented blue color, and darker blue found in stoma, veins and root caps in 5d seedlings. Further, hypocotyls of 14 d seedings was in blue too. No blue colors were observed in ProABI5::GUS or ProABF4::GUS transgenic seedlings.(3) Seed germination and seedling growth of Arabidopsis thaliana were inihibited by different concentrations of ABA, Na Cl, sorbitol and sucrose, respectively. The inhibition is stronger as the concentrations go higher.(4). The effects of ABA, drought, cold and high salt on m RNA contents of the ABI5 subfamily members: The m RNA contents of 14 d Arabidopsis thaliana seedlings treated by 20μM ABA for 0h, 0.5h, 1h, 2h, 4h and 6h, were increased over treatment time for ABF1, ABF2, ABF3, ABF4 and ABI5, which increased significantly for ABF1 and ABI5. The m RNA contents for DPBF2, DPBF3, DPBF4 were not observed obvious regular changes. The m RNA contents for ABF1, ABF4 ABI5, DPBF3 and DPBF4 increased over treatment time in 14 d old Arabidopsis thaliana seedlings treated by 100 m M sorbitol. m RNA for ABF1 and ABI5 increased largely, but regular changes of m RNA for ABF2, ABF3 and DPBF2 were not observed. m RNAs for ABF1 and ABI5 were accumulated significantly over treatment time in 14 d old Arabidopsis thaliana seedlings treated under 4℃. Obvious changes of m RNA contents were not observed over treatment in the 14 d old Arabidopsis thaliana seedings treated by 100 m M Na Cl.