RNAi沉默淀粉分支酶基因SBEI对玉米直链淀粉合成的影响

淀粉分支酶(SBE)是淀粉合成的限速酶。为了研究SBEI沉默对直链淀粉合成的影响,克隆了玉米(Zeamays)淀粉分支酶SBEI基因片段,构建了SBEI的RNAi表达载体pBAC418,用基因枪将其导入玉米自交系幼胚愈伤组织,经木糖筛选获得了7株转化再生植株。利用FAD2 intron和xylA基因探针对T0代再生玉米植株进行DNAdotblot和PCR-Southern检测,证实5株为阳性植株,其中4株正常结实。SBEI基因沉默对阳性再生玉米株系籽粒的含油量没有显著影响;蛋白质含量显著高于受体对照;总淀粉含量与对照相比无显著差异,转基因株系直链淀粉含量平均提高了9.8%。

Increasing Amylose Content by Silencing SBEI Using RNA Interference in Maize

Starch branching enzyme (SBE) plays an important role in the synthesis of starch. We used RNA interference (RNAi) to silence the SBE gene (SBEI) to research the effect of SBEI silencing in maize. The xylA gene from Escherichia coli encoding xylose isomerase was used as a selective marker. We constructed the expression vector pBAC418 containing the CaMV 35S promotor driving xylA and the reverse repeat sequence of SBEI driven by its corresponding promotor with biolistic PDS1000/He to bombard maize calli induced from the elite inbred lines. The selection was made on media containing 50% concentrations of xylose. We obtained 7 transformed plantlets. Gene integration was confirmed in 5 transformed plantlets by DNA dot blot analysis and PCR-Southern hybridization of xylA and FAD2 intron. Total protein, starch, oil and amylose contents of regenerated plants were detected. Oil content of regenerated plants showed no significant change, the total protein content was increased, and starch content was decreased a little. Mean amylose content increased 9.8% in transgenic maize lines.