Construction of Yeast Two-hybrid Bait Vector and the Screening of Proteins Interacting with HbICE1 in Hevea brasiliensis

Cold stress is a major issue in the rubber plantation areas of China. It not only affects rubber production, but also threatens the survival of the rubber tree. ICE1 is a transcriptional activator in the cold-response pathway of Arabidopsis thaliana L.. However, most key genes, including HbICE1, in the cold-response pathways of Hevea brasiliensis (Willd. ex A. Juss.) Muell. have not yet been cloned, and the proteins that interact with HbICE1 are unclear, which has hampered the advance of molecular mechanism research of cold tolerance in H. brasiliensis. We cloned HbICE1 and constructed its bait vector, and then transformed pGBKT-7-HbICE1 into yeast Y2H strain. The transformed yeast cells were plated on selection medium for bait auto-activation test. The auto-activation of HbICE1 was inhibited on selection medium (SD/-Trp/-His/-Ade) supplemented with 30 mmol/L 3-AT (3-amino-1,2,4-triazole) and higher concentrations. Thereafter, the proteins that interacted with HbICE1 were screened through the yeast two-hybrid method, which included DNA-binding protein, ribosomal protein and function unknown proteins. This study provides an important theoretical basis for the molecular mechanism of the rubber tree’s response to cold stress.