pGCS Vector Kit
(Kit # 1000000107 )

Depositing Lab:   Hai-Ning Du

As a multipurpose expression vector, pCS2+ backbone-based expression plasmids are widely used for high-level expression of messenger RNAs (mRNAs) or proteins in mammalian/avian culture cells or Xenopus/zebrafish embryos.

This kit contains a series of pCS2+ backbone-based Gateway destination vectors (pGCS), bearing either amino- or carboxyl-terminal tags, including Myc, HA, Flag, GST and eGFP epitopes, which allow the generation of various tagged proteins in a large variety of vertebrate organisms.

This kit will be sent as individual bacterial stabs.

Original Publication

Construction of a series of pCS2+ backbone-based Gateway vectors for overexpressing various tagged proteins in vertebrates. Wang HY, Li Y, Xue T, Cheng N, Du HN. Acta Biochim Biophys Sin (Shanghai). 2016 Dec;48(12):1128-1134. PubMed PMID: 27797719.

Description

The pGCS Vector Kit consists of pCS2+ backbone-based expression plasmids that are compatible for high level transient expression in multiple organisms, including mammalian/avian culture cells, and Xenopus/zebrafish embryos.

The pGCS Gateway destination vectors featured in this kit bear either amino- or carboxyl-terminal tags, including Myc, HA, Flag, GST and eGFP epitopes, offering an easy cut and paste approach for the generation of various tagged proteins. The pGCS vectors possess all features of pCS2+, as well as the core cassette of Gateway destination vectors for recombination reactions.

Schematic representation of vector maps and core elements of pGCS Gateway destination cloning vectors.
The pGCS vectors possess all features of pCS2+ vector, as well as the core cassette of Gateway destination vector for recombination reaction. The core cassette contains two attR sites (marked with dark blue) flanking bacterial ‘death’ gene (ccdB, marked with brown) and chloramphenicol resistance gene (CmR, marked with pea green). Various tags are labeled with light green.

Protocols

A standard LR reaction was performed according to the Invitrogen Gateway Technology Manual with some modifications.

• An aliquot of 1 μl (25 ng/μl) of an Entry clone containing any verified gene was mixed with 1 μl of the same concentration of either pGCS destination vector.
• Then 0.5 μl of LR clonase enzyme mix was added and vortexed.
• After at least 1 h of incubation at room temperature, proteinase K was added to terminate the reaction.
• An aliquot of 1 μl of the reaction mixture was transformed into DH5α competent cells.
• Positive clones were selected on LB agar plates with ampicillin antibiotics.

How to Cite this Kit

These plasmids were created by your colleagues. Please acknowledge the Principal Investigator, cite the article in which they were created, and include Addgene in the Materials and Methods of your future publications.

For your Materials and Methods section:

“The pGCS Vector Kit was a gift from Hai-Ning Du (Addgene kit #1000000107).”

For your Reference section:

Construction of a series of pCS2+ backbone-based Gateway vectors for overexpressing various tagged proteins in vertebrates. Wang HY, Li Y, Xue T, Cheng N, Du HN. Acta Biochim Biophys Sin (Shanghai). 2016 Dec;48(12):1128-1134. PubMed PMID: 27797719.

Kit Contents

The pGCS Vector Kit contains 12 plasmids. Please refer to the individual plasmid pages below for more details on each plasmid in this kit:

ID	Name
85718	pGCS-N1(6xMYC)
85719	pGCS-N2(3xHA)
85720	pGCS-N3(3xFLAG)
85722	pGCS-N5(GST)
85723	pGCS-N6(eGFP)
85724	pGCS-C1m(5xMYC)
85725	pGCS-C2m(3xHA)
85726	pGCS-C3m(3xFLAG)
85728	pGCS-C5(GST)
85729	pGCS-C6(eGFP)
85730	pGCS1
85731	pGCS2