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Cloning and Agrobacterium-Mediated Transformation of Phosphate-Starvation Tolerant Related MicroRNAs and Their Targets into Soybean

Author: SunDianChen
Tutor: ZhouXinAn
School: Chinese Academy of Agricultural Sciences
Course: Crop Genetics and Breeding
Keywords: microRNA399 microRNA398 Target genes Low Phosphorus Stress
CLC: S565.1
Type: Master's thesis
Year: 2011
Downloads: 124
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Abstract


Soybeans are one of the world's most important oil crops . High total phosphorus content of soil , but most can not be absorbed by crops use , in addition to the large number of application of phosphate fertilizer must exacerbate environmental pollution . Soybean low phosphorus stress tolerated , protect the environment and soybean high yield , stable yield has important practical significance . Mining -related gene using genetic engineering methods , and target gene into soybean through genetic transformation is modified soybean low phosphorus stress tolerance is an important way . In recent years , microRNA and its target gene 's mechanism of action become a hot research . microRNA is a class 21-25nt endogenous non-coding RNA , they inhibit the expression of target genes through transcriptional or post-transcriptional level . microRNA399 and microRNA398 two Arabidopsis phosphorus stress resistance microRNA and be able to respond quickly in low phosphorus conditions . This experiment was constructed the two plant Arabidopsis pre-microRNA399b and soybean microRNA398a target gene GmSODC expression vector , and verify that the function of these genes induced by Agrobacterium rhizogenes transformed roots by Agrobacterium tumefaciens -mediated transformation of soybean cotyledon node transgenic plant . The main findings are as follows : 1, by PCR method amplified from Arabidopsis and soybean to of two purposes of genes (microRNA399b GmSODC ) LIC (Ligation-Independent cloning) method to connect to the target gene binary plant expression vector the pJG045 will verify plasmid using the freeze-thaw method imported into Agrobacterium EHA105 construct two plant expression vector . 2, the desired gene are respectively connected to the plants with the GFP expression vector pCX-DG , using freeze-thaw method is introduced into the Agrobacterium rhizogenes K599 to soybean cotyledons section receptor , gene function of the two purposes of the preliminary verification. The low phosphorus hydroponic experiments , GmSODC improvement soybean tolerance to low phosphorus stress ability , not obvious microRNA399b . Agrobacterium-mediated cotyledon node wears by grafting combination of conventional PCR and fluorescence quantitative PCR validation obtain a the conversion microRNA399b T0 plants obtained 13 the conversion GmSODC T0 plants , four T1 plants .

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CLC: > Agricultural Sciences > Crop > Economic crops > Oil crops > Soybean
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