Total RNA was extracted from hippocampus in mice using TRIzol reagent (Takara, Japan). And then the cDNA was synthesized by Prime-Script™ RT regent Kits (Takara, Japan). The mRNA expression levels were measured using qRT-PCR by SYBR Premix Ex Taq (Takara, Japan). The critical threshold cycle (Ct) value was determined and the relative quantification data were calculated with the 2-ΔΔCt method, the GAPDH was served as a reference. The primers were as follows: IL-1β-forward: 5′-GAGTCTGCACAGTTCCCCAA-3′, IL-1β-reverse: 5′-TGTCCCGACCATTGCTGTTT-3′; TNF-α-forward: 5′-CGTC AGCCGATTTGCCATTT-3′, TNF-α-reverse: 5′-CTCCCTCAGGGGTGTCCTTA-3′; TGF-β-forward: 5′-GAACTCGCTTTGTCTCCA-3′, TGF-β-reverse: 5′-TACAGTCGCAGTATAACCTCA-3′; IL-10-forward: 5′-TCTCCGAGATGCCTTCAGCAGA-3′, IL-10-reverse: 5′-TCAGACAAGGCTTGGCAACCCA-3′; GAPDH-forward: 5′-ATGGGGAAGGTGAAGGT-3′, GAPDH-reverse: 5′-AAGCTTCCCGTTCTCAG-3′.
Prime script rt regent kits
Manufactured by Takara Bio
Sourced in Japan
The Prime-Script™ RT regent Kits are reverse transcription reagent kits designed for the synthesis of first-strand cDNA from RNA templates. The kits contain the necessary components, including a reverse transcriptase enzyme, to perform this fundamental step in gene expression analysis and other RNA-based applications.
Automatically generated - may contain errors
Lab products found in correlation
Trizol reagent, by Takara Bio (2 mentions)
Nanodrop 2000, by Thermo Fisher Scientific (2 mentions)
Sybr premix ex taq, by Takara Bio (1 mentions)
Cfx connect rt qpcr system, by Bio-Rad (1 mentions)
Sybr premix ex taq 2 kit, by Takara Bio (1 mentions)
Trizol rna isolation system, by Thermo Fisher Scientific (1 mentions)
Power sybr green pcr master mix, by Takara Bio (1 mentions)
3 protocols using prime script rt regent kits
1
Investigating inflammatory gene expression in mouse hippocampus
2
Quantifying mRNA Expression in AML Cells
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Total RNA from AML cells was isolated using TRIzol reagent (#9108, Takara, Japan), then reverse transcribed into cDNA with PrimeScript RT regent Kits (#RR600, Takara, Japan). RT-qPCR was carried out with SYBR® Premix Ex Taq™ II kits (#RR820A, Takara) in a CFX Connect™ RT-qPCR System (Bio-Rad, USA). All primers were synthesised by Sangon Biotech (Shanghai, China). Primer sequences are given in Table S1 . The number of cycles at which the fluorescence intensity increment reaches a threshold in each sample tube was taken as the Ct value. The Ct values of the gene to be tested and the internal reference gene were read separately for each specimen. Using β-actin as the internal reference, the relative expression of the gene to be tested was further calculated for each specimen using 2-ΔΔCt23 (link).
3
Quantitative Analysis of Autophagy Genes
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Total RNA samples were extracted using a TRIzol RNA isolation system (Invitrogen). Purity and concentration of mRNA samples were measured using a NanoDrop 2000 spectrophotometer (Thermo Fisher Scientific, Inc., Wilmington, DE, USA). Samples were reverse transcribed (RT) into cDNAs using PrimeScript™ RT regent kits (Takara Biotechnology, Shiga, Japan). cDNAs were used for RT-qPCR analysis using a Power SYBR Green PCR Master Mix (Takara Biotechnology). qPCR was performed with 2 µl aliquots of cDNA (10 ng/µl) using specific primer pairs. Primers were as follows: LC3A forward, 5′-GACCGCTGTAAGGAGGTGC-3′ and reverse, 5′-CTTGACCAACTCGCTCATGTTA-3′; LC3B forward, 5′-TTATAGAGCGATACAAGGGGGAG-3′ and reverse, 5′-CGCCGTCTGATTATCTTGATGAG-3′; GAPDH forward, 5′-AGGTCGGTGTGAACGGATTTG-3′ and reverse, 5′-TGTAGACCATGTAGTTGAGGTCA-3′. Samples were amplified by 95°C for 2 min, 40 cycles of 95°C for 30 sec, 95°C for 5 sec, 60°C for 5 sec, and a 72°C for 10 min. Data were calculated using the 2−ΔΔCq method (13 (link)).
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