RT was performed by using iScript Reverse Transcription Supermix kit (Bio-Rad). PCR reactions were run on a CFX96 Touch Real Time PCR (Bio-Rad) with SsoAdvanced™ Universal SYBR® Green Supermix (Bio-Rad). Primers for qPCR (Cldn2, Dlx1, Dlx5, Etv5, Gabrb1, Gabrg2, Gabra3, Gad2, Gapdh, Itgb3, Lhx1, Lhx6, Mef2c, Nkx2.1, Nkx2.2, Nrp1, VegfA, Wnt10a, Wnt10b) were obtained from Thermo Fisher Scientific. The housekeeping gene Gapdh was used as a reference. The relative gene expression among different samples and subsequent fold increase in Vgatfl/fl versus VgatECKO endothelial cells or GABAergic neurons was determined according to published methodology44 (link).
Wnt10b
Manufactured by Thermo Fisher Scientific
Wnt10b is a recombinant protein produced in human cells. It is a member of the Wnt family of secreted signaling proteins involved in various cellular processes.
Automatically generated - may contain errors
Lab products found in correlation
Ssoadvanced universal sybr green supermix, by Bio-Rad (1 mentions)
Vegf a, by Thermo Fisher Scientific (1 mentions)
Gapdh, by Thermo Fisher Scientific (1 mentions)
Cfx96 touch real time pcr, by Bio-Rad (1 mentions)
Iscript reverse transcription supermix kit, by Bio-Rad (1 mentions)
Cldn2, by Thermo Fisher Scientific (1 mentions)
Gabrb1, by Thermo Fisher Scientific (1 mentions)
Itgb3, by Thermo Fisher Scientific (1 mentions)
Mef2c, by Thermo Fisher Scientific (1 mentions)
Gapdh, by Abcam (1 mentions)
Phenyl methyl sulfonyl fluoride (pmsf), by Cell Signaling Technology (1 mentions)
β catenin, by BD (1 mentions)
2 protocols using wnt10b
1
Quantitative Analysis of Gene Expression
2
Western Blot Analysis of Wnt Signaling
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Cells were lysed with an appropriate amount of Nonidet‐P40 Cell Lysis Buffer (20 mM Tris HCl, 137 mM NaCl, 10% glycerol, 1% nonidet‐P40, 2 mM EDTA) supplemented with complete protease inhibitor and 1 mM PMSF (Cell Signaling Technology) with 1× Phosphatase Inhibitor Cocktail (Cell Signaling Technology). Proteins separated by 10% and 12% SDS gel‐electrophoreses were transferred to a polyvinylidene difluoride (PVDF) membrane by semi‐dry blotting. The membrane was incubated with primary antibodies against WNT10B (ThermoFisher), β‐catenin (BD Transduction Laboratories), ABC‐Active β‐catenin (Merk Millipore), FZD4 (Abcam), FZD5 (Abcam), FZD6 (ThermoFisher Scientific), GAPDH (Abcam) at 4°C for 2 h (Table S3 ).
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