Clonging ABP1 Gene And Primary Research On Transformation In Super Rice

The super rice "Liangyoupeijiu" was used as material in this paper.The cDNA of Auxin-binding protein 1 (ABP1) have been cloned, and the GeneBinary expression recombinant of ABP1 have been constructed. We havescreened the better condition of the cellar culture of the super rice invitro, and we have transformed the ABP1 gene into cellar of the super riceby Agrobacteria-mediated transformation. The details as following:1. The cloning of the ABP1 cDNA in super riceA rice cDNA sequence was searched from nucleotide sequence database;the authors analyzed its full-length sequence by bioinformatics andlaboratory bench-work. The results suggest the sequence is rice ABP1 cDNA.Based the sequence, ABP1 cDNA of super rice was amplified by RT-PCR andcloned for sequencing; the results of sequencing were marched with theprovided sequence by BLAST.2. The construction of the expression vector of ABP1 geneAccording to the cDNA sequence of ABP1 gene, two specific primerswhich contiains the restriction endonuclease site of upstream primer XbaI or ABP just downstream primer BamH I rescpectively were designed andthe sequence of GGCG for the protection bases. After PCR, the PCR productsdigested by XbaⅠand BamHⅠwas ligateded with the vector pBI121 whichdigested by the same enzymes. And then the liagtion products weretransformed into E. coli JM109 for the construction of the expressionrecombinant of ABP1 gene.3. The optimization screening of the better conditions of the cellarculture of the super rice in vitro.a. By change of the condition of the super rice tissue culture, weconfirmed that the best induced media component and concentration: MS+sugrose 30 g/L+Agar 7 g/L+2,4-D 2 mg/L+KT 1 mg/L; the best differentiation media component and concentration: MS+sugrose 30 g/L+Agar 7 g/L+6-BA 2mg/L+KT 1mg/L+NAA 0.5 mg/L。b. The effects of the cellar differentiation of super rice by theaddition Cu element into differentiation media4. Agrobacteria mediated transformation of ABP1 gene.The ABP1 gene fused in expression vector pBI121 was transformed into thecellar of the super rice by Agrobacteria-mediated transformation. Byscreening of the transformed cellar with antibiotic, and detecting of thetransgenic cellar by PCR and GUS staining, the results showed thatthe target gene was inserted into the genome of the super rice.