pcDNA/PGK-neo* and pcDNA-Redβ/PGK-neo* have been described (Zhang et al., 2003 (link)). A derivative of the latter, pcDNA-nlsRedβ/PGK-neo*, was made by adding AGGATCCAACC ATG GGC CCT AAA AAG AAG CGT AAA GTC GCC AGT in place of AGGAATTCACC ATG AGT at the N-terminus of Redβ. To make pTRE-Tight-Redβ, the nls-Redβ open-reading-frame was removed from pcDNA-nlsRedβ/PGK-neo* as an 825 bp BamHI/HindIII fragment and cloned into the multiple cloning site of pTRE-Tight (BD Biosciences, USA). Oligonucleotides were commercially synthesised by Sigma-Aldrich Inc.
Ptre tight
Manufactured by BD
Sourced in United States
The PTRE-Tight is a laboratory equipment designed for maintaining a tight seal. Its core function is to provide a secure and controlled environment for various experimental setups.
Automatically generated - may contain errors
Lab products found in correlation
2 protocols using ptre tight
1
Constructing Inducible Red Fluorescent Protein
2
Inducible Expression of KCTD17 and Trichoplein in RPE1 Cells
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Tet-On RPE1 cell lines that expressed myc-KCTD17 or MBP-trichoplein-flag (WT or K50/57R) were established with the same procedure described previously29 (link)46 (link)47 (link). The rtTA-advanced segment and the tTS transcriptional silencer segment from pTet-On Advanced and pQC-tTS-IN (BD Clontech) were recombined into the retroviral vector pDEST-PQCXIP and pDEST-PQCXIN, respectively, by the LR reaction (Invitrogen) to generate PQCXIN-Tet-On ADV and PQCXIP-tTS. The Elongation factor 1 alpha promoter (EF) in CSII-EF-MCS (a gift from Hiroyuki Miyoshi, RIKEN BioResource Center, Tsukuba, Japan) was replaced with a Tet-responsive promoter (TRE-Tight) from pTRE-Tight (BD Clontech) followed by a modified RfA fragment (Invitrogen) to make a Tet-responsive lentivirus vector, CSII-TRE-Tight-RfA. Fusion cDNAs with siRNA-resistant KCTD17 and trichoplein were recombined into the lentiviral vector by the LR reaction (Invitrogen) to generate CSII-TRE-Tight-myc-KCTD17 and CSII-TRE-Tight-MBP-trichoplein-3xFLAG, respectively. For induction of myc-KCTD17 or MBP-trichoplein-flag, Tet-On RPE1 cells were treated with 30 or 100 ng ml−1 doxycycline (Sigma-Aldrich), respectively.
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