The reverse transcripts were performed using an Invitrogen SuperScript Reagent Kit. The primer designed using the Oligo6 software. For RT-PCR, the SYBR® Premix Ex Taq™ (TAKARA) was used on a Bio-Rad CFX96 real-time PCR detection system (Bio-Rad, Hercules, CA). Gene expression was analyzed for samples at 6, 12, 24, 48 and 72 hours post-inoculation of R and S genotypes. All reactions for each gene were performed in triplicate. The relative expression level of each gene among samples was calculated using the 2-△△Ct method with normalization to the internal reference actin gene. The parameters of thermal cycle were 95°C for 30 s, followed by 40 cycles of 95°C for 10 s, 50–56°C for 25 s at a volume of 20 μl.
Superscript reagent kit
Manufactured by Thermo Fisher Scientific
Sourced in United States
The SuperScript Reagent Kit is a set of reagents designed for reverse transcription, which is the process of synthesizing complementary DNA (cDNA) from messenger RNA (mRNA) templates. The kit contains the necessary components, including a reverse transcriptase enzyme, buffer, and other essential reagents, to facilitate the conversion of mRNA into cDNA for downstream applications.
Automatically generated - may contain errors
Lab products found in correlation
Sybr premix ex taq, by Takara Bio (4 mentions)
Cfx96 real time pcr detection system, by Bio-Rad (2 mentions)
Iq5 qrt pcr detection system, by Bio-Rad (1 mentions)
Iq5 real time pcr detection system, by Bio-Rad (1 mentions)
Dreamtaq dna polymerase, by Thermo Fisher Scientific (1 mentions)
Gel doc ez imager, by Bio-Rad (1 mentions)
6 protocols using superscript reagent kit
1
Real-Time PCR Analysis of Gene Expression
2
Single-cell RT-PCR of Labeled Cells
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Single‐cell RT‐PCR was performed as described previously.[11 , 48 ] GFP‐labeled microglial cell and astrocytes were extracted from Cx3cr1Gfp mice and Aldh1l1Gfp mice, respectively. The contents of neurons are harvested from the lamina II of spinal cord slices from C57Bl/6 mice. Reverse transcriptions were performed using an Invitrogen SuperScript Reagent Kit (Invitrogen, CA, USA). The first step of the reverse transcription system was performed at 37 °C for 40 min and then at 80 °C for 10 min to remove the genomic DNA. The second reaction was then performed at 50 °C for 50 min and 70 °C for 15 min. Single‐cell PCR was performed as a nested PCR. The external primers were used for the primary amplification, and internal primers were used for the following secondary amplification. The sequences of nested PCR primers are depicted in Table S3 of the Supporting Information. All PCR products were visualized using agarose gel (2.5%) electrophoresis with Gel‐Red.
3
Gene Expression Analysis in Pods
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Reverse transcriptions were performed using an Invitrogen SuperScript Reagent Kit. We performed qRT-PCR using a Bio-Rad IQ5 qRT-PCR detection system (Bio-Rad, Hercules, CA, USA) and the SYBR® Premix ExTaq® (TAKARA). Gene expression of both lines (pw and WT) was detected for pod samples at DAF 20, DAF 40, and DAF 60, and three repeats of each reaction for individual genes were performed. The relative expression of each gene among different samples was calculated by using the 2−ΔΔCt method and normalized by using the internal reference actin gene. Thermal cycle parameters were 95°C for 30 s, followed by 40 cycles of 95°C for 10 s, and between 50 and 56°C for 25 s in a 20 μl volume.
4
Real-Time qRT-PCR Analysis of Gene Expression
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The reverse transcriptions were performed using an Invitrogen SuperScript Reagent Kit. The primer was designed using the Oligo6 software. For RT-PCR, the SYBR® Premix ExTaq™ (TAKARA) was used on a Bio-Rad IQ5 real-time PCR detection system (Bio-Rad, Hercules, CA). Gene expression was analyzed for samples at 20, 40, and 60 DAF of WT and mutant. All reactions for each gene were performed in triplicate. The relative expression level of each gene among samples was calculated using the 2−ΔΔCt method with normalization to the internal reference actin gene. The parameters of thermal cycle were 95°C for 30 s, followed by 40 cycles of 95°C for 10 s and 50–56°C for 25 s at a volume of 20 μl.
5
Peanut Gene Expression Analysis under Salt Stress
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The reverse transcripts were carried out with an Invitrogen SuperScript Reagent Kit. The primer was designed by Oligo6 software. For RT-qPCR, the SYBR® Premix Ex Taq™ (TAKARA) was adopted on a Bio-Rad CFX96 real-time PCR detection system (Bio-Rad, Hercules, CA). Gene expression of the samples of S2 and S4 at 6, 12, 24 and 48 h after the salinity stress was analyzed. The relative expression level of each gene among the samples was computed with the 2-ΔΔCt method with normalization to the internal reference genes Actin and TUA5 of peanut, which has been reported to have stable expression in leaves when subjected to salt stress treatment [11 (link)]. The relative expression level of each gene in roots at 6, 12, 24 and 48 h after the salt stress had also been analyzed and confirmed, with normalization to the internal reference genes Actin and HDC of peanut, which has been reported to have stable expression in roots when subjected to salt stress treatment [11 (link)]. All reactions of each gene were conducted with three biological replicates respectively. The parameters of thermal cycle were 95°C for 30 s, followed by 40 cycles of 95°C for 10 s, 50–56°C for 25 s with the volume being 20 μl.
6
Cardiac Gene Expression Analysis
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Heart tissues were homogenized in Tri-reagent and RNA was precipitated using isopropanol. RNA pellets were washed with 75% ethanol and then dissolved in RNA storage solution. Reverse transcription was performed using the Superscript reagent kit (Invitrogen, 18 080-044) to obtain cDNA. PCR was performed using Dream Taq DNA polymerase (Thermo Fisher Scientific EP0701) and customized PCR primers (Supplementary Material, Table S6 ), and PCR products were separated on 5% agarose gels. Images were acquired using Gel Doc EZ imager (BIO-RAD) and quantified with ImageJ image processing software.
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!