Cloning And Agrobacterium-Mediated Transformation Of Phosphate-Starvation Tolerant Related MicroRNAs And Their Targets Into Soybean

Soybean is one of the most important oil crops in the world. Phosphorus is one of the most important mineral elements for plant growth and the least available essential nutrients in the soil . The use of fertilizers, are costly to farmers and the environment.The improvement of the ability to adapt low-phosphate stress of soybean and other crops is very urgent. Plant genetic transformation is an important way to improve low-phosphate tolerability for soybean.In recent years, the mechanism of interaction between miRNA and its target gene becomes the focus of research. MiRNAs are a class of small, non-coding RNAs, with 21-25 nucleotide length. These small RNAs induce transcriptional or posttranscriptional gene silencing by guiding heterochromatin formation, mRNA degradation, or translational inhibition. MiR399 and miR398 involved in phosphorus starvation response. Two plant expression vectors pre-miR399b and the target of miR398a were constructed in this experiment , and Agrobacterium tumefaciens–mediated soybean cotyledonary node system was used for soybean genetic transformation,the main research results were as follows:1.MiR399 and GmSODC(target of miR398)were cloned and inserted into the binary vector pJG045.Two plant expression vectors were constructed and transformed into EHA105 successfully.2.The two genes were also inserted into the multiple cloning sites of the binary vector pCX-DG with GFP.The two genes'function was tested by Agrobacterium rhizogenes-mediated soybean cotyledonary node system preliminarily. Results show that GmSODC performed expected function, but microRNA399 did not.3.Through PCR and qRT-PCR, one T0 plant with miR399, thirteen T0 plants and four T1 plants with GmSODC gave positive results.