BioVector® pSEVA-gRic6T Plasmid假单胞菌基因组编辑质粒指导RNA递送和供体模板质粒BioVector NTCC保藏中心

价　　格：¥49950
货　　号：BioVector® pSEVA-gRic6T
产　　地：北京

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Product Name / 产品名称:
BioVector® pSEVA-gRic6T Plasmid
BioVector® pSEVA-gRic6T质粒

Description / 描述:
The BioVector® pSEVA-gRic6T plasmid is an E. coli-Pseudomonas putida KT2440 shuttle vector designed as a guide RNA (sgRNA) delivery and donor template plasmid for CRISPR-based genome editing in pseudomonads. It contains a synthetic sgRNA expression cassette targeting the P. putida KT2440 genome, along with homologous arms for homology-directed repair, enabling precise genomic modifications. This plasmid is part of a two-plasmid CRISPR system for efficient genome editing in Pseudomonas species.
BioVector® pSEVA-gRic6T 质粒是一种大肠杆菌-恶臭假单胞菌 KT2440穿梭载体，设计用于在假单胞菌中实现基于CRISPR的基因组编辑，作为指导RNA递送和供体模板质粒。它包含一个靶向恶臭假单胞菌 KT2440基因组的合成sgRNA表达盒，以及用于同源定向修复的同源臂，能够实现精确的基因组修饰。该质粒是双质粒CRISPR系统的一部分，用于在假单胞菌属中进行高效的基因组编辑。

Depositing Lab / 保藏实验室:
Sheng Yang Lab, Center for Excellence in Molecular Plant Sciences (CEMPS), CAS / 杨晟实验室，中国科学院分子植物科学卓越创新中心

Plasmid Type / 质粒类型:

E. coli-Pseudomonas shuttle vector / 大肠杆菌-假单胞菌穿梭载体
sgRNA delivery plasmid / sgRNA递送质粒
Donor template plasmid (with homologous arms) / 供体模板质粒（含同源臂）
CRISPR editing component plasmid / CRISPR编辑组件质粒

Species / 物种:

Escherichia coli (for propagation) / 大肠杆菌（用于扩增）
Pseudomonas putida KT2440 and related pseudomonads (for application) / 恶臭假单胞菌KT2440及相关假单胞菌（用于应用）

Vector Backbone / 载体骨架:
pSEVA644 + pTargetF (hybrid backbone) / pSEVA644 + pTargetF杂合骨架

Backbone Size (without insert) / 骨架大小（不含插入片段）:
~4500 bp / 约4500 bp

Total Vector Size / 总载体大小:
~4600 bp (estimated, based on backbone + insert) / 约4600 bp（根据骨架+插入片段估算）

Key Components / 关键组件:

sgRNA expression cassette / sgRNA表达盒: Contains synthetic guide RNA targeting P. putida KT2440 genome
pj23119 promoter / pj23119启动子: Constitutive synthetic promoter for sgRNA expression
Homologous arms / 同源臂: Flanking sequences for homology-directed repair (specific to target locus) / 用于同源定向修复的侧翼序列
Broad-host-range origin / 广宿主范围复制起点: For maintenance in Pseudomonas species

Gene/Insert Details / 基因/插入片段详情:

Parameter / 参数	Specification / 规格
Gene/Insert Name / 基因名称	sgRNA (targeting P. putida KT2440 genome) / 靶向恶臭假单胞菌KT2440基因组的sgRNA
Species / 物种	Synthetic / 合成
Insert Size / 插入片段大小	96 bp
Promoter / 启动子	pj23119 (synthetic constitutive promoter) / pj23119（合成组成型启动子）
Target Gene / 靶基因	nicC (PP_3944) and others as designed / nicC (PP_3944) 及其他设计靶点

Growth Conditions in Bacteria / 细菌生长条件:

Parameter / 参数	Specification / 规格
Bacterial Resistance / 细菌抗性	BioVector® Gentamicin (10 μg/mL for E. coli; 50 μg/mL for Pseudomonas)
Growth Strain / 生长菌株	DH5α or similar cloning strains
Growth Temperature / 培养温度	37°C
Copy Number / 拷贝数	Low to moderate / 低至中等

Applications / 应用:

CRISPR Genome Editing: Used in conjunction with Cas9-expressing plasmid (e.g., pCAS-RK2K/T) for targeted gene deletions, insertions, and replacements in P. putida KT2440
Gene Deletion: Successfully used to delete nicC gene (1149 bp) with 100% efficiency
Gene Insertion: Enables insertion of heterologous genes such as rhla (from P. aeruginosa), dCas9, and T7 RNA polymerase into specific genomic loci
Metabolic Engineering: Facilitates genetic modification of Pseudomonas for bioproduction and synthetic biology applications
Multiplex Editing: Part of a versatile CRISPR toolkit for various genome modifications in pseudomonads

Experimental Performance / 实验性能:
The pSEVA-gRic6T plasmid has demonstrated high efficiency in various genome editing applications when used in P. putida KT2440 harboring a Cas9-expressing plasmid:
pSEVA-gRic6T质粒在携带Cas9表达质粒的恶臭假单胞菌KT2440中展现出高效的基因组编辑性能：

Application / 应用	Target / 靶点	Efficiency / 效率
Gene deletion / 基因敲除	nicC (1149 bp)	15/15 (100%)
Gene replacement / 基因替换	nicC::rhla (888 bp insert)	38/40 (95%)
Gene insertion / 基因插入	nicC::dCas9 (4107 bp)	20/20 (100%)
Gene insertion / 基因插入	nicC::T7 RNAP (4518 bp)	14/20 (70%)

Related Plasmids / 相关质粒:

Product Name / 产品名称	Description / 描述
BioVector® pCAS-RK2K/T Plasmid	Cas9-expressing plasmid for use with pSEVA-gRic6T in two-plasmid CRISPR system 与pSEVA-gRic6T配合使用的Cas9表达质粒
BioVector® pSEVA-gRic5T Plasmid	Alternative sgRNA plasmid targeting nicC with different efficiency 靶向nicC的替代sgRNA质粒，效率不同
BioVector® pSEVA-gRic6PAM2 Plasmid	For single nucleotide editing at PAM sites 用于PAM位点的单核苷酸编辑
BioVector® pSEVA-gRxxxxT Series	Plasmids targeting various loci (PP_0552, PP_3361, PP_3733, etc.) 靶向不同位点的系列质粒

Quality Control / 质量控制:
Plasmid is verified by restriction digestion and sequencing. Full sequence information is available from the depositor.
质粒经过限制性酶切和测序验证。完整序列信息可向保藏者获取。

Storage / 存储:

Plasmid is sent as a bacterial stab in agar or as purified DNA. Upon receipt, streak onto a plate with BioVector® Gentamicin (10 μg/mL), grow a single colony, and prepare a glycerol stock for long-term storage at -80°C.
质粒以细菌穿刺菌或纯化DNA形式提供。收到后，应在含 BioVector® 庆大霉素 (10 μg/mL) 的平板上划线，挑取单克隆，并制备甘油菌于-80°C长期保存。

Citation / 引用:
When using this plasmid, please cite the following article:
使用此质粒时，请引用以下文献：

Sun J, Wang Q, Jiang Y, Wen Z, Yang L, Wu J, Yang S. Genome editing and transcriptional repression in Pseudomonas putida KT2440 via the type II CRISPR system. Microb Cell Fact. 2018 Mar 17;17(1):41.

Important Note / 重要提示:

The pSEVA-gRic6T plasmid is designed for use in a two-plasmid CRISPR system and requires co-transformation with a Cas9-expressing plasmid (e.g., pCAS-RK2K or pCAS-RK2T) for genome editing applications.
pSEVA-gRic6T质粒设计用于双质粒CRISPR系统，进行基因组编辑时需要与Cas9表达质粒（如pCAS-RK2K或pCAS-RK2T）共转化。
The plasmid contains homologous arms specific to the target locus; for editing different genes, the homologous arms need to be redesigned and cloned.
该质粒含有靶向特定位点的同源臂；如需编辑不同基因，需重新设计和克隆相应的同源臂。
Gentamicin resistance is functional in both E. coli and Pseudomonas hosts at appropriate concentrations (10 μg/mL for E. coli, 50 μg/mL for Pseudomonas).
庆大霉素抗性在大肠杆菌和假单胞菌宿主中均具有功能，需使用适当浓度（大肠杆菌用10 μg/mL，假单胞菌用50 μg/mL）。