Factors affecting the transformation of 'Marshall McIntosh' apple by Agrobacterium tumefaciens

被引：39

作者：

Bolar J.P. ^{[1
]}

Brown S.K. ^{[1
]}

Norelli J.L. ^{[2
]}

Aldwinckle H.S. ^{[2
]}

机构：

[1] Department of Horticultural Sciences, Cornell University, New York State Agric. Exp. Station, Geneva

[2] Department of Plant Pathology, Cornell University, New York State Agric. Exp. Station, Geneva

来源：

Plant Cell, Tissue and Organ Culture | 1998年 / 55卷 / 1期

关键词：

Agar/Gelrite combination; Aminoglycosides; Malus x domestica Borkh; Marshall McIntosh; Neomycin phosphotransferase; β-glucuronidase;

D O I：

10.1023/A:1026414031217

中图分类号：

学科分类号：

摘要：

The goal of this research was to develop an efficient transformation system for 'Marshall McIntosh' apple. To determine the optimum combination of agar and Gelrite gelling agents in the media to maximize regeneration and minimize hyperhydicity (vitrification), the following combinations of agar (A)+Gelrite (G) in g l-1 were tested: 7.0 A+0 G; 5.2 A+0.6 G; 3.5 A+2.5 G; 1.7 A+1.8 G; and 0 A+2.5 G. Both 5.2 A+0.6 G and 3.5 A+1.2 G provided greatest regeneration of healthy non-hyperhydric shoots. To determine the optimal concentration of aminoglycoside for the selection and regeneration of transgenic 'Marshall McIntosh' on agar-Gelrite-based media, kanamycin was tested at 0, 10, 25, 50, 75 and 100 mg l-1, and paromomycin was tested at 0, 50, 100, 150, 200 and 250 mg l-1. Kanamycin was more effective than paromomycin in the initial selection of transgenics. For selection of transformants of 'Marshall McIntosh', the use of kanamycin at 25 mg l-1 on 5.2 A+0.6 G solidified medium is suggested. By optimizing the medium and selection conditions, a protocol was developed that resulted in four transgenic lines as confirmed by a GUS assay, NPT II ELISA, PCR, and Southern analysis. In repeated experiments with this protocol, transformation efficiencies of 3.1 and 2.6% were obtained.

引用

页码：31 / 38

页数：7

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