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以蕺菜DNA为模板,采用正交试验设计,对影响蕺菜rDNA ITS区扩增的重要参数进行了优化试验,以期建立蕺菜rDNA ITS扩增反应的优化体系。结果表明,最优蕺菜ITS-PCR的反应体系为25μl反应体系中含Taq酶0.75 U、Mg~(2+)+(1 mmol/L)1.0μl、dNTP(0.15 mmol/L)1.5μl、引物对(0.6μmol/L)1.2μl、模板DNA(10 ng/μl)2.0μl、10×buffer(不含Mg~(2+))2.5μl。这一体系的建立为利用蕺菜rDNA ITS区研究蕺菜种质资源的系统进化提供了标准化程序。
Using chives DNA as a template and using orthogonal design, the optimized parameters affecting the amplification of the rDNA ITS region of the leek were optimized to establish an optimization system for ITS amplification of rDNA from Amaranthus. The results showed that the optimal reaction system for ITS-PCR of leeks was 25 μl of reaction system containing 0.75 U of Taq enzyme, 1.0 μl of Mg 2+ + (1 mmol/L), and 1.5 μl of dNTP (0.15 mmol/L). Primer pair (0.6 μmol/L) 1.2 μl, template DNA (10 ng/μl) 2.0 μl, 10×buffer (without Mg 2+ ) 2.5 μl. The establishment of this system provides a standardized procedure for the systematic evolution of Amaranth germplasm resources using the rDNA ITS region of Amaranthus.