Sybr green qpcr master mix reagent system

Rat mammary gland carcinoma walker 256 cells in the logarithmic growth phase were plated in 24-well plates at 1 × 10⁵ cells, and infected with PBS, LV containing shMRPS23 or LV carrying shCtrl. Total RNA was extracted from cells and tissues using the Trizol reagent (Invitrogen, Carlsbad, CA, USA) and reverse transcribed PCR using the PrimeScript™ RT Master Mix (Takara, Ohtsu, Japan). Expression levels of targeted genes were analysed by qRT-PCR on a SYBR Green qPCR Master Mix reagent system (Takara, Ohtsu, Japan). The PCR primers used are listed as follows: MRPS23: Forward 5′-TGTCAGCGGTTTGT-GGAG-3′; Reverse 5′-CAGT CTTTGCTTCTCTTACTCGTC-3′; β-actin: Forward 5′-GG AGATTACTGCCCTGGCTCCTA-3′; Reverse 5′-GACTC ATCGTACTCCTGCTTG-CTG-3′; p53: Forward 5′-GTC GGCTCCGACTATACCACTATC-3′; Reverse 5′-CTCTC TTTGCACT-CCCTGGGGG-3′; p21^WAF1/CIP1: Forward 5′-AG TATGCCGTCGTCTGTTCG-3′; Reverse 5′-TCAAAGTT CCACCGTTCTCG-3′; Cyt c: Forward 5′-AAAGGAG GCAAGCATAAGACTG-3′; Reverse 5′- TGTTCTTGTT GGCATCTGTGT-3′; E-cadherin: Forward 5′-TCTCT TGTCCCTTCCA-CAGC-3′; Reverse 5′-CTCCAGACC CACACCAAAGT-3′; Vimentin: Forward 5′-GGATTT CTCTG-CCTCTTCCA-3′; Reverse 5′-CACCTGTCCGT CTCTGGTTT-3′.

On day 21 after corresponding treatments, the Vglut2-cre^+/– mice were deeply anesthetized with 2% pentobarbital sodium (0.5 mL/100 g, intraperitoneal injection) and the L4–L6 spinal cord was immediately transferred to an ice-chilled lysis buffer through laminectomy. Total RNA was extracted from the spinal cord using Trizol reagent and reverse transcribed according to the manufacturer’s instructions (Takara, Tokyo, Japan). The expression of target genes was analyzed using the ViiA7 Dx system (Applied Biosystems, Carlsbad, CA, United States), with the SYBR Green qPCR Master Mix reagent system (Takara). The forward and reverse primers used in this study were: