pRI101-EGFP植物基因转化双元荧光表达载体-BioVector NTCC典型培养物保藏中心

The BioVector™ pRI101-EGFP plasmid is a specialized expression vector used for genetic transformation in plants. It's a combination of the pRI101 plasmid backbone and the Enhanced Green Fluorescent Protein (EGFP) gene. The plasmid is designed to introduce a foreign gene into plant cells using Agrobacterium-mediated transformation, with the EGFP gene serving as a reporter.

Vector Features & Function

The pRI101-EGFP plasmid has a specific set of features that make it suitable for its purpose:

• Plant-Specific Components: It contains a strong constitutive promoter, the CaMV 35S promoter, to drive high-level expression of the EGFP gene in plant cells. It also includes the 5'-UTR of an alcohol dehydrogenase (ADH) gene from either Arabidopsis thaliana (in pRI101-AN) or rice (in pRI101-ON), which acts as a translational enhancer to boost protein production.

• Reporter Gene (EGFP): The EGFP gene is cloned into the plasmid's multiple cloning site. When the plasmid is successfully delivered into a plant cell, the EGFP protein is expressed. This allows researchers to easily and visually identify which cells have been successfully transformed by observing the green fluorescence under a microscope.

• Binary Vector: pRI101 is a binary vector, meaning it can replicate in both E. coli and Agrobacterium tumefaciens. This is essential for the transformation process:

  • It is first cloned and propagated in E. coli.

  • Then, it's transferred into Agrobacterium, which acts as the delivery vehicle to insert the T-DNA (which contains the EGFP gene) into the plant's genome.

• Selection Markers: The plasmid contains genes for antibiotic resistance, such as kanamycin resistance (NeoR/KanR), to select for bacteria and plant cells that have successfully taken up the plasmid.

Applications

This plasmid is a fundamental tool for plant genetic engineering, specifically for:

• Verifying Transformation: Its primary use is to visually confirm successful genetic transformation in plants without needing more complex, destructive methods.

• Studying Gene Expression: Researchers can use the EGFP fluorescence to track the expression patterns and subcellular localization of a gene of interest by creating a fusion protein with EGFP.

• High-Throughput Screening: It enables high-throughput screening of a large number of transformed plants or cells, as fluorescence is a fast and simple readout.

• Plant-Specific Gene Function Studies: The pRI101 vector backbone is optimized for stable integration into the plant genome, allowing for long-term studies of gene function.

BioVector™ pRI101-EGFP质粒是一种用于植物基因转化的专用表达载体，它结合了 pRI101 质粒骨架和增强型绿色荧光蛋白（EGFP）基因。该质粒旨在利用农杆菌介导的转化方法将外源基因导入植物细胞，其中 EGFP 基因作为报告基因。

载体特征与功能

pRI101-EGFP 质粒具有一套使其适用于该目的的特定功能：

• 植物特异性组件: 它包含一个强力的组成型启动子——CaMV 35S 启动子，用于在植物细胞中驱动 EGFP 基因的高水平表达。它还包含来自拟南芥（在 pRI101-AN 中）或水稻（在 pRI101-ON 中）的乙醇脱氢酶（ADH）5' 非翻译区，该区域可作为翻译增强子来提高蛋白质产量。

• 报告基因（EGFP）: EGFP 基因被克隆到质粒的多克隆位点中。当质粒成功导入植物细胞后，EGFP 蛋白会被表达。这使得研究人员能够通过显微镜下观察绿色荧光，轻松直观地识别哪些细胞已成功转化。

• 二元载体: pRI101 是一种二元载体，这意味着它可以在大肠杆菌和根癌农杆菌中复制。这对于转化过程至关重要：

  • 它首先在大肠杆菌中进行克隆和扩增。

  • 然后，它被转移到农杆菌中，农杆菌充当递送载体，将 T-DNA（包含 EGFP 基因）插入植物的基因组中。

• 选择标记: 质粒含有抗生素抗性基因，例如卡那霉素抗性基因（NeoR/KanR），用于筛选那些已成功吸收质粒的细菌和植物细胞。

应用

该质粒是植物基因工程的基础工具，特别用于以下方面：

• 验证转化: 其主要用途是直观地确认植物的成功基因转化，而无需使用更复杂、具有破坏性的方法。

• 研究基因表达: 研究人员可以通过将目的基因与 EGFP 融合，利用 EGFP 荧光来追踪其表达模式和亚细胞定位。

• 高通量筛选: 由于荧光是一种快速简单的读数，因此它能够对大量转化的植物或细胞进行高通量筛选。

• 植物特异性基因功能研究: pRI101 载体骨架经过优化，可以稳定地整合到植物基因组中，从而可以对基因功能进行长期研究。

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