通过微粒轰击实现枣椰树的遗传转化

Genetic Transformation of Date Palm Via Microprojectile Bombardment.

作者信息

Mousavi Mousa, Mousavi Amir, Habashi Ali A, Arzani Kazem, Dehsara Bahareh, Brajeh Mohsen

机构信息

Department of Horticulture Science, Faculty of Agriculture, Shahid Chamran University, Ahvaz (Ahwaz), Iran.

National Institute of Genetic Engineering and Biotechnology (NIGEB), Tehran, Iran.

出版信息

Methods Mol Biol. 2017;1637:269-280. doi: 10.1007/978-1-4939-7156-5_22.

DOI:10.1007/978-1-4939-7156-5_22

PMID:28755352

Abstract

Efficient protocols for date palm embryogenic callus and somatic embryo transformation with uidA gene are described in this chapter. The embryogenic callus transformation procedure is 1.6 μm gold particle size coated with 2.5 μg DNA (pAct1-D plasmid), 1100 psi helium pressure, 9 cm target distance, 26 inHg vacuum pressure, 3 mm distance between the rupture disk and macrocarrier, and osmotic pretreatment with 0.4 M mannitol followed by 60 min air desiccation. The somatic embryo transformation procedure is 0.6 μm gold particle size coated with 2.5 μg DNA (pAct1-D plasmid), 1350 psi helium pressure, 6 cm target distance, 28 inHg vacuum pressure, 3 mm distance between the rupture disk and macrocarrier, and osmotic pretreatment with 0.4 M mannitol followed by 60 min air desiccation. Protocols for analysis of the transgenic plantlets have also been described.

摘要

本章介绍了利用uidA基因对枣椰胚性愈伤组织和体细胞胚进行转化的高效方案。胚性愈伤组织转化程序为：使用粒径1.6μm的金颗粒，包被2.5μg DNA（pAct1 - D质粒），氦气压力1100psi，靶距离9cm，真空压力26inHg，破裂盘与宏载体之间距离3mm，先用0.4M甘露醇进行渗透预处理，然后空气干燥60分钟。体细胞胚转化程序为：使用粒径0.6μm的金颗粒，包被2.5μg DNA（pAct1 - D质粒），氦气压力1350psi，靶距离6cm，真空压力28inHg，破裂盘与宏载体之间距离3mm，先用0.4M甘露醇进行渗透预处理，然后空气干燥60分钟。还描述了转基因植株的分析方案。