Product Name / 產(chǎn)品名稱:
BioVector? pSEVA-gRic6T Plasmid
BioVector? pSEVA-gRic6T質(zhì)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 質(zhì)粒是一種大腸桿菌-惡臭假單胞菌 KT2440穿梭載體，設(shè)計用于在假單胞菌中實現(xiàn)基于CRISPR的基因組編輯，作為指導(dǎo)RNA遞送和供體模板質(zhì)粒。它包含一個靶向惡臭假單胞菌 KT2440基因組的合成sgRNA表達盒，以及用于同源定向修復(fù)的同源臂，能夠?qū)崿F(xiàn)精確的基因組修飾。該質(zhì)粒是雙質(zhì)粒CRISPR系統(tǒng)的一部分，用于在假單胞菌屬中進行高效的基因組編輯。

Depositing Lab / 保藏實驗室:
Sheng Yang Lab, Center for Excellence in Molecular Plant Sciences (CEMPS), CAS / 楊晟實驗室，中國科學(xué)院分子植物科學(xué)卓越創(chuàng)新中心

Plasmid Type / 質(zhì)粒類型:

• E. coli-Pseudomonas shuttle vector / 大腸桿菌-假單胞菌穿梭載體

• sgRNA delivery plasmid / sgRNA遞送質(zhì)粒

• Donor template plasmid (with homologous arms) / 供體模板質(zhì)粒（含同源臂）

• CRISPR editing component plasmid / CRISPR編輯組件質(zhì)粒

Species / 物種:

• Escherichia coli (for propagation) / 大腸桿菌（用于擴增）

• Pseudomonas putida KT2440 and related pseudomonads (for application) / 惡臭假單胞菌KT2440及相關(guān)假單胞菌（用于應(yīng)用）

Vector Backbone / 載體骨架:
pSEVA644 + pTargetF (hybrid backbone) / pSEVA644 + pTargetF雜合骨架

Backbone Size (without insert) / 骨架大?。ú缓迦肫危?
~4500 bp / 約4500 bp

Total Vector Size / 總載體大小:
~4600 bp (estimated, based on backbone + insert) / 約4600 bp（根據(jù)骨架+插入片段估算）

Key Components / 關(guān)鍵組件:

• sgRNA expression cassette / sgRNA表達盒: Contains synthetic guide RNA targeting P. putida KT2440 genome

• pj23119 promoter / pj23119啟動子: Constitutive synthetic promoter for sgRNA expression

• Homologous arms / 同源臂: Flanking sequences for homology-directed repair (specific to target locus) / 用于同源定向修復(fù)的側(cè)翼序列

• Broad-host-range origin / 廣宿主范圍復(fù)制起點: For maintenance in Pseudomonas species

Gene/Insert Details / 基因/插入片段詳情:

Growth Conditions in Bacteria / 細菌生長條件:

Applications / 應(yīng)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 / 實驗性能:
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質(zhì)粒在攜帶Cas9表達質(zhì)粒的惡臭假單胞菌KT2440中展現(xiàn)出高效的基因組編輯性能：

Related Plasmids / 相關(guān)質(zhì)粒:

Quality Control / 質(zhì)量控制:
Plasmid is verified by restriction digestion and sequencing. Full sequence information is available from the depositor.
質(zhì)粒經(jīng)過限制性酶切和測序驗證。完整序列信息可向保藏者獲取。

Storage / 存儲:

• 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.

• 質(zhì)粒以細菌穿刺菌或純化DNA形式提供。收到后，應(yīng)在含 BioVector? 慶大霉素 (10 μg/mL) 的平板上劃線，挑取單克隆，并制備甘油菌于-80°C長期保存。

Citation / 引用:
When using this plasmid, please cite the following article:
使用此質(zhì)粒時，請引用以下文獻：

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質(zhì)粒設(shè)計用于雙質(zhì)粒CRISPR系統(tǒng)，進行基因組編輯時需要與Cas9表達質(zhì)粒（如pCAS-RK2K或pCAS-RK2T）共轉(zhuǎn)化。

• The plasmid contains homologous arms specific to the target locus; for editing different genes, the homologous arms need to be redesigned and cloned.

• 該質(zhì)粒含有靶向特定位點的同源臂；如需編輯不同基因，需重新設(shè)計和克隆相應(yīng)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).

• 慶大霉素抗性在大腸桿菌和假單胞菌宿主中均具有功能，需使用適當濃度（大腸桿菌用10 μg/mL，假單胞菌用50 μg/mL）。

BioVector? Gentamicin
BioVector? 慶大霉素

For selection of pSEVA-gRic6T plasmid and its derivatives in bacterial hosts.
用于在細菌宿主中選擇pSEVA-gRic6T質(zhì)粒及其衍生質(zhì)粒。

BioVector NTCC質(zhì)粒載體菌株細胞蛋白抗體基因保藏中心

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