Author: Jacob Peter Matson; Amy M. House; Gavin D. Grant; Huaitong Wu; Joanna Perez; Jeanette Gowen Cook
Title: Intrinsic checkpoint deficiency during cell cycle re-entry from quiescence Document date: 2019_2_22
ID: dsbucda9_38
Snippet: To make the RPE1 line containing PCNA-mTurq2, Cdc6-mVenus, and DHB-mCherry, the plasmids were transfected into 293T with pCI-GPZ or ΔNRF and VSVG virus packaging plasmids with 50 ug/mL Polyethylenimine (Aldrich Chemistry). Viral supernatants were transduced onto RPE1 cells with 8 ug/mL Polybrene (Millipore). A clonal cell line was picked based on fluorescence of all 3 biosensors. siRNA transfections and drug treatment siRNA concentration and seq.....
Document: To make the RPE1 line containing PCNA-mTurq2, Cdc6-mVenus, and DHB-mCherry, the plasmids were transfected into 293T with pCI-GPZ or ΔNRF and VSVG virus packaging plasmids with 50 ug/mL Polyethylenimine (Aldrich Chemistry). Viral supernatants were transduced onto RPE1 cells with 8 ug/mL Polybrene (Millipore). A clonal cell line was picked based on fluorescence of all 3 biosensors. siRNA transfections and drug treatment siRNA concentration and sequences: siControl, (siLuciferase) 100 nM (synthesized by Life Technologies) 5′-CUUACGCUGAGUACUUCGA-3′ siCdt1 A, mixture of 4 sequences, 25 nM each. (siGENOME CDT1 siRNA, Dharmacon) 5'-CCAAGGAGGCACAGAAGCA-3' 5'-GCUUCAACGUGGAUGAAGU-3' 5'-UCUCCGGGCCAGAAGAUAA-3' 5'-GGACAUGAUGCGUAGGCGU-3' siCdt1 B, 50 nM (synthesized by Life Technologies)
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