柚类、橘类叶肉原生质体分离方法研究

Protoplast Isolation from Pummelo and Tangerine Leaves

原生质体是植物遗传改良的重要实验体系之一,而原生质体的分离是该体系的基础.对柑橘叶肉原生质体来说,柠檬、橙类等品种的叶肉原生质体比较容易分离,而柚类、橘类等柑橘品种则相对困难.本研究采用椪柑、黄岩本地早、沙田柚、冰糖橘4个品种为试材,枳壳试管苗为对照,系统研究了不同基因型、不同叶龄、不同酶种类、不同酶液组合对叶肉原生质体分离的影响,探讨了影响柚类、橘类等柑橘叶肉原生质体分离的因素,从中找出了各品种较适宜的分离条件.在适宜条件下,4种供试品种的原生质体得率(FW)可达10×106～20×106 g-1.

Protoplast is an important system for plant genetic improvement, and the isolation of protoplast is an essential step for the protoplast marnipulation. Among different varieties of citrus, it has been confirmed that isolations were fairly accessible from the leaves of lemon and navel orange, but difficult from pummelo and tangerine. In this paper, using Ponkan （Citrus reticulata）, Huangyanbendizao tangerine （C. succosa Hort Ex Tanaka）, Shatian pummelo （C. grandis）, Bingtang tangerine （C. reticulata） as study material, and Poncirus trifoliate as control, the effects of genus, age of seedlings, enzyme type, enzyme combination on the leaf protoplast isolation of pummelo and tangerine were studied, and the suitable conditions for the leaf protoplast isolation of the four studied varieties were optimized. The best results of 10× 10^6-20 × 10^6 g^-1 protoplast were obtained with the optimized conditions.