Pcdh cmv mcs ef1 cogfp vector

Manufactured by System Biosciences

Sourced in United States

The PCDH-CMV-MCS-EF1-coGFP vector is a plasmid designed for the expression of a gene of interest. It contains a CMV promoter for driving expression, a multiple cloning site for insertion of the gene, and an EF1 promoter-driven codon-optimized GFP reporter.

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Lab products found in correlation

Polybrene, by Merck Group (2 mentions) Psicheck 2 vector, by Promega (1 mentions) Pspax2, by Addgene (1 mentions) Jetprime, by Polyplus Transfection (1 mentions) Pmd2 g, by Addgene (1 mentions) Plko 1, by Addgene (1 mentions)

Spelling variants of this product

PCDH vector (30 citations) PCDH-CMV-MCS-EF1-coGFP (2 citations) PCDH-CMV-coGFP (2 citations)

3 protocols using pcdh cmv mcs ef1 cogfp vector

Cloning of Abhd2 and miR-485 Constructs

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CDS sequence of Abhd2 was amplified from the genomic DNA of ES cells by PCR. The PCR primers were as follows: forward primer: 5′-CGGAATTCGCCACCATGGACTACAAAGACGATGACGACAAGATGAATGCCATGCTAGAGACCC-3′ (underlined letters indicate an EcoRI restriction site) and reverse primer: 5′-ATAAGAATGCGGCCGCTCATTCCAACTCGGCCTCCATCTGCTCC-3′ (underlined letters indicate a NotI restriction site). The PCR sample was ligated into the pcDNA 3.1. a fragment carrying pre-miR-485 was amplified from genomic DNA of J1 ESCs and cloned into the pCDH-CMV-MCS-EF1-coGFP vector (System Biosciences, Mountain View, CA, USA). The PCR primers were as follows: forward primer: 5′-GCTCTAGACTACCACAGGAGCTTCCAGAATA-3′ (underlined letters indicate a XbaI restriction site) and reverse primer: 5′-ATAAGAATGCGGCCGCTAGCTTGGACACTGGGATAACTG-3′ (underlined letters indicate a NotI restriction site).
The 3′-UTR of genes that contain putative miRNA binding sites were amplified from J1 ESC cDNA through PCR and then cloned into psiCHECK-2 Vector (Promega) for Luciferase assays. All primer sequences used were shown in the Supplementary Table (Table S2)

Yu M., Zhang L., Liu Y., Liu D, & Guo Z. (2019). Retinoic Acid Induces Differentiation of Mouse F9 Embryonic Carcinoma Cell by Modulating the miR-485 Targeting of Abhd2. International Journal of Molecular Sciences, 20(9), 2071.

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Knockdown and Overexpression of Key Genes in hASCs and hBMSCs

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The knockdown of NOVA1, DNAJC10 and NCOR2 in hASCs or hBMSCs was performed by using the lentivirus short hairpin RNA (shRNA) knockdown vector system PLKO.1 (Addgene, USA). Two NOVA1 shRNA sequences were designed as follows, A, 5′-CCGGCAGACCACCGTTAATCCAGATCTCGAGATCTGGATTAACGGTGGTCTGTTTTTG-3′; B, 5′- CCGGACCAAGTCCTCTCCATCTGATCTCGAGATCAGATGGAGAGGACTTGGTTTTTTG-3′. The DNAJC10 shRNA sequence is 5′-CCGGGCACCAGACATCTGTAGTAATCTCGAGATTACTACAGATGTCTGGTGCTTTTTG-3′. The NCOR2 shRNA sequence is 5′-CCGGATATGACCAGTGGGAAGAGTCCTCGAGGACTCTTCCCACTGGTCATATTTTTTG-3′. The scramble sequence 5′-CCTAAGGTTAAGTCGCCCTCGCTCGAGCGAGGGCGACTTAACCTTAGG-3′ was taken as a control.
For gene overexpression, the full coding sequence (CDS) of NOVA1 or DNAJC10 was inserted into the pCDH-CMV-MCS-EF1-CoGFP vector (System Biosciences, Mountain View, CA, USA), and named pCDH-NOVA1 and pCDH-DNAJC10, respectively. The empty pCDH-CMV-MCS-EF1-CoGFP vector served as a control. For viral packaging, the shRNA and overexpression vectors were transfected into 293T cells together with psPAX2 and pMD2.G (Addgene, USA) with jetPRIME (Polyplus-transfection, USA). The supernatants containing the virus were collected and concentrated before infecting cells in the presence of 5 mg/ml polybrene (Millipore, USA).

Yang Z., Dong P., Cao J., Lin N., Ma S., Cao R., Cai L., Wang L., Cao C., Xue Y., Pan J., Li X., Wang K., Liu Q., Li C., Gong F., Fu X, & Xiao R. (2023). NOVA1 prevents overactivation of the unfolded protein response and facilitates chromatin access during human white adipogenesis. Nucleic Acids Research, 51(13), 6981-6998.

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Lentiviral Transduction of TRIM32

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The coding sequences of TRIM32 were cloned into the pCDH‐CMV‐MCS‐EF1‐coGFP vector (System Biosciences, Mountain View, CA, USA). TRIM32 short hairpin RNA (shRNA) (target sequence: 5′‐GGUGGAAAGCUUUGGUGUU‐3′) was cloned into the pLKO.1 vector. Lentiviral particle production was performed according to other reports.14 Cells were infected with recombinant lentivirus‐transducing units plus 5 mg/mL Polybrene (Sigma, St Louis, MO, USA).

Wang C., Xu J., Fu H., Zhang Y., Zhang X., Yang D., Zhu Z., Wei Z., Hu Z., Yan R, & Cai Q. (2018). TRIM32 promotes cell proliferation and invasion by activating β‐catenin signalling in gastric cancer. Journal of Cellular and Molecular Medicine, 22(10), 5020-5028.

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