rnai consortium trcn0000240480 trcn0000181026 pljc6 3xha tmem192 addgene  (Addgene inc)

Journal: bioRxiv

Article Title: Modelling oxaliplatin resistance in colorectal cancer reveals a SERPINE1 -based gene signature (RESIST-M) and therapeutic strategies for pro-metastatic CMS4 subtype

doi: 10.1101/2024.12.17.628817

Figure Lengend Snippet: Immunoblotting of PAI-1 protein in HCT116 MD and HD knocked down for the protein using two unique shRNAs ( A ). Fold change of PAI-1 protein in cell culture supernatant of HCT116 MD and HD knocked down for the protein relative to MD shRNA control (shCtrl) ( B ). Fold change of dose response to 48 hrs oxaliplatin treatment in HCT116 MD and HD knocked down for the protein relative to MD shCtrl ( C ). Representative dose response curves of HCT116-MD shCtrl, PAI-1 sh3 and sh5 ( D ) and HCT116-HD shCtrl, PAI-1 sh3 and sh5 ( E ) to 48 hrs of oxaliplatin treatment. F-F’, Inhibition of metastasis in oxaliplatin treated HCT116 model using tiplaxtinin. Three representative H&E staining and corresponding quantifications ( F and F’ ) of lung sections from mice harboring HCT116 PAR xenografts treated with oxaliplatin, alone or in combination with tiplaxtinin, until spontaneous metastasis (refer to section ‘Impact of concomitant administration of tiplaxtinin and oxaliplatin on metastasis’ under Materials and Methods). For quantification number of nodule positive area was normalized to total lung area (n= at least 8). Statistical significance was determined using Ordinary one-way ANOVA followed by Sidak’s multiple comparisons test for Fig. B, C, and using two-tailed unpaired t-test for Fig. F’. A p-value of <0.05 was considered significant for all analyses, unless stated otherwise.

Article Snippet: To generate stable knockdown lines, suitable bacteria clones from the RNAi Consortium (TRC) shRNA Library (Broad Institute) were selected and sub-cultured in terrific Broth (CUS-4051-1L, Axil Scientific) supplemented with 1X carbenicillin (10177012, ThermoFisher).

Techniques: Western Blot, Cell Culture, shRNA, Control, Inhibition, Staining, Two Tailed Test