Screening Xylem-Specific Expression Gene And PtoZF Transfermation Into Populus Tomentosa

Populus tomentosa is widely planted in northern China and used in Paper industry. It has the characteristics of excellent material and resistance. Xylem is the transport organization of vascular plants, and also has the function of mechanical support. Compared with the researches of the key enzyme gene in the xylem, there are fewer researches in the area of the specific promoter element and transcription factor.Constructing a yeast one-hybrid system and screening binding proteins of specific regulatory elements from Populus tomentosa were conducted to research the transcription regulation network in xylem. To provide references for researching timber synthesis related transcription factor, we constructed three copies of ABRE-element、AC-element and W-element into bait vector, then established and screened yeast one-hybrid library by using AC-element as bait with the Populus tomentosa as the plant material.In this study, we selected the PtoZF gene, specifically expressing in xylem, as the research object. The PtoZF-overexpressed/antiexpressed vectors were transformed into Populus tomentosa by agrobacterium-mediated transformation. It provided base on the research of the transcription factors function, and made a great significance of understanding the molecular mechanisms of xylem formation, and genetic engineering breeding of improving the properties.The detail experiment results are as follows:(1)Using the GUS gene as PCR-template, amplified it with its upstream including respectively ABRE-box、AC-box and W-box in the 5’end, together with its downstream primer. Construct the PCR products into pAbAi vector and then remove the GUS fragments.(2) We constructed yeast one-hybrid library and screened 1.3×106 clones for AC-box inserted yeast strains without the interruption of rRNA.5 binding proteins were detected. Analyze their possible function by blasting on NCB1.(3)It was confirmed that the best concentration of Kanamycin for the transformed Populous was 15 mg/L. We have obtained the transgenic plant regeneration from Populustomentos.