The Cloning And Rice Transformation Of The Ferritin Gene From Adzuki Bean (Vigna Angularis)

Through technique of rapid amplification cDNA ends (RACE), cDNA of adzuki bean ferritin gene was cloned.To improve the iron content of rice, we transferred the entire coding sequence of the adzuki bean ferritin gene into rice by particle gun transformation, which not only helped to breed new variety of transgenic rice with ferritin, but also solve the problem of dietary iron deficiency. The main results and significances are as follows:1, According to homology sequence from soybean, pea and so on, by means of designing degenerate primer, we gained ferritin gene cDNA from adzuki bean with 3'-RACE and 5'-RACE techniques. The ferritin gene cDNA is 1030bp long with an open reading frame of 768bp encoding for a protein of 255 amino acids. The access number in Genbank is AY663197.2 By means of taking measures of reclaiming DNA with big molecular weight which come from reverse transcript before the circularity of ligase enzyme.problems of recombination among small segments were setteled. The probability of successful amplification was greatly improved.With this way, technical supports were supplied for RACE.3 In order to transfer rice and study ferritin gene expression in rice, we constructed the plant expression vector-the pCambial301-UbiN-AFerCDS, which is an expression vector that take the ubiquitin to carry.4 Through the approach of particle gun transformation, the expression vector with ferritin gene was transformed into upland rice by PCR analysis.5 The iron level in leaf of transgenic plants varies from 144.1 to 240.0 μ g/gDW. Given that the iron content in leaf of nontransgenic plants was 110.6μg/gDW, we estimated an increase of 2.17 times in iron content of leaf from the transgenic upland rice lines.