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以蓖麻茎秆为试材,研究了影响cDNA-AFLP标记蓖麻差异基因反应体系的几个关键因素,建立了适宜蓖麻的cDNA-AFLP分析体系。结果表明:当裂解液为1.4 mL时提取的RNA浓度最高,EcoRⅠ和MseⅠ的组合6 h可将ds-cDNA酶切完全,与相应接头连接过夜后产物用于预扩增,预扩增产物稀释15倍时作为选择性扩增模板可以获得条带丰富且重复性好的结果。蓖麻cDNA-AFLP反应体系的建立为进一步研究蓖麻株高、叶型等相关基因奠定了基础。
Using castor stem as test material, we studied several key factors that affect the cDNA-AFLP-labeled ricin differential gene reaction system and established a cDNA-AFLP analysis system for castor. The results showed that when the concentration of lysate was 1.4 mL, the concentration of RNA extracted was the highest. The combination of EcoRI and Mse I could complete the ds-cDNA digestion for 6 h. After ligation with the corresponding linker overnight, the product was used for pre-amplification. 15 times as a selective amplification template can get rich and reproducible band results. The establishment of ricin cDNA-AFLP reaction system lays the foundation for further study on plant height, leaf type and other related genes in castor.