橡胶树细胞悬浮系电激转化体系的初步研究  

Transformation of Hevea Suspension Cells by Electroporation

作　　者：肖成江[1,2] 程汉[2] 祝建顺[1,2] 安泽伟[2] 黄华孙[2] 

机构地区：[1]海南大学农学院,海南海口570208 [2]中国热带农业科学院橡胶研究所国家橡胶树育种中心,海南儋州571737

出　　处：《热带作物学报》2011年第2期245-250,共6页Chinese Journal of Tropical Crops

基　　金：国家天然橡胶产业技术体系(CARS-34);中央级公益性科研院所基本科研业务费专项基金项目(No.XJSYWFZX2009-03)

摘　　要：以橡胶树属种间杂交系B1(H.brasiliensis×H.nitida)的细胞悬浮系为试材,采用均匀设计结合偏最小二乘(PLS)回归,以瞬时表达率(Y1)、细胞成活率(Y2)为试验指标综合评价,以因变量最大值为优化方向,同时对两个试验指标进行回归建模。实验结果表明,正常接种后培养3 d对数期的悬浮细胞,在10μF、2 kn、1 300 V/cm的电激条件下,蔗糖浓度为0.4 mol/L的电激转化液,加入终浓度为90μg/mL的线性质粒CaMV35S-EGFP-NOS,600 mbar压强下抽真空1 min,可使细胞瞬时表达率达到0.648 1%,成活率达到56.984 7%。通过对电激转移中相关因素的优化,建立了一种适用于橡胶树悬浮细胞系较稳定的电激转移体系,为橡胶树悬浮细胞系转基因的研究提供有效方法。The uniform design coupled with Partial Least-squares（PLS）regression synthetical was utilized to evaluate the comprehensive test indices as transformation efficiency（Y1）and cell survival rate（Y2）by analyzing Hevea Rubber Hybrid B1（H.brasiliensis×H.nitida）suspension cells.Regression modeling of two test indices were built to optimize the maximum value of the dependent variable.The results showed that under the electric shock conditions of 10 μF,2 kn,1 300 V/cm,a final concentration of 90 μg/mL linear plasmid CaMV35S-EGFP-NOS in 0.4 mol/L sucrose electroporation solution and one minute of 600 mbar vacuum-pumping,logarithmic phase suspended cells,cultured 3 d,could reach an instant expression efficiency and survival rate in 0.648 1% and 56.984 7%,respectively.This study established a system that was applicable to a stable electric shock of Hevea rubber suspension cells and provided an effective method for the research of Hevea rubber suspension cells transgene by optimizing some correlative factors during electroporation.

关 键 词：橡胶树 细胞悬浮系 均匀设计 偏最小二乘回归 

分 类 号：S794.1[农业科学—林木遗传育种]