英文文献阅读报告.ppt

Reading Report
Agro bacterium
农杆菌是生活在植物根的表面依靠由根组织渗透出
来的营养物质生存的一类普遍存在于土壤中的革兰
氏阴性细菌。
农杆菌是一种天然的植物遗传转化体系,被誉为“自
然界最小的遗传工程师”。可以通过将目的基因插入
到经过改造的T-DNA区,借助农杆菌的感染实现外
源基因向植物细胞的转移和整合,然后通过细胞和
组织培养技术,得到转基因植物。
Agro bacterium rhizogenes-mediated transformation provides a system for rapid and efficient transformation of plant tissues.
The influence of several factors
plant genotype
A. rhizogenes culture stage
A. rhizogenes cell titre
co-culture period of A. rhizogenes
acetosyringone concentration
a routine, efficient and genotype-independent method of inducing hairy roots by A. rhizogenes in peanuts was established.
诱导农杆菌Vir基因活化,从而促进外源基因的整合
Plant material
Seven peanut genotypes including Luhua11,Huayu16,Huayu28, Baisha1016, Yuanhua8, Xinhua1, and Xinhua5 were used to assess the effect of plant genotype on -mediated transformation.
Agrobacterium rhizogenes strain and plasmid
pCAMBIA2300
The vector pGFP-GUSPlus which contained two highly-expressed reporter genes, GFP gene and GUS gene.
Induction of roots by microinjection
Peanut hairy-root induction
A-5-day old seedling for microinjection;
B-microinjection of A. rhizogenes on hypocotyls at different sides;
C-Tumor(肿瘤) emerging 1 week after microinjection;
D-Induced roots appearing 2 weeks after inoculation;
E-Transformed root system
Detection of GFP or synthetic cry8Ea1 gene in transgenic roots by RT-PCR
Total RNA from induced roots was isolated by the TRIzol method and treated with RNase-free DNase according to the manufacturer’s instructions.
One microliter of first strand cDNA was used as template for PCR amplification.
The two sets of primer pairs used in the assay were primers VirHF/VirHR, primer GFPF (base pairs 76–93),50-CTTCTCGTTGGGGTCTTT-30, and primer GFPR (base pairs 627–644), 50-G-30. Primer jc8EF2, 50-GGCGGCAGCAT TCAAACTCAA-30, and primer jc8ER2,-30 were used to detect the transcription of the synthetic cry8Ea1 gene(暗黑鳃金龟抗性基因).
Histochemical assay for GUS activity
Root segments