Syber green

Manufactured by Merck Group

Sourced in India

Syber Green is a fluorescent dye used in molecular biology for the detection and quantification of DNA. It binds to double-stranded DNA and emits a green fluorescent signal upon excitation. The dye is commonly used in applications such as real-time PCR, gel electrophoresis, and DNA sequencing to visualize and analyze DNA samples.

Automatically generated - may contain errors

Lab products found in correlation

Nanodrop nd 1000, by Thermo Fisher Scientific (1 mentions) Light cycler equipment, by Roche (1 mentions) Rneasy mini kit, by Qiagen (1 mentions) Rnase inhibitor, by Thermo Fisher Scientific (1 mentions) Superscript 3 reverse transcriptase, by Thermo Fisher Scientific (1 mentions) Steponeplus system, by Thermo Fisher Scientific (1 mentions) Viia7 real time pcr machine, by Thermo Fisher Scientific (1 mentions) Universal cdna synthesis kit, by Qiagen (1 mentions)

3 protocols using syber green

Quantitative real-time PCR protocol

Check if the same lab product or an alternative is used in the 5 most similar protocols

RNA was extracted using metal beads in a Tissue Lyser and the RNeasy Mini Kit (Qiagen) following the instructions provided by the supplier. Quantity and quality of RNA were analyzed using Nanodrop ND-1000 (Thermo Scientific).
RT-PCRs were performed in the Genomic Facility at CBMSO using Light Cycler equipment (Roche) and primers labeled with Syber Green (Supplementary Table 1) (Sigma).
A geometric mean of three stably expressed and commonly used reference genes (ywhaz, gapdh and 18s) was used for data normalization in the liver and a geometric mean of two (ppia and b2m) was used for data normalization in WAT and BAT. qPCRs and statistical analysis of the data were performed using GenEx software.

Vila-Bedmar R., Cruces-Sande M., Lucas E., Willemen H.L., Heijnen C.J., Kavelaars A., Mayor F J.r, & Murga C. (2015). Reversal of diet-induced obesity and insulin resistance by inducible genetic ablation of GRK2. Science signaling, 8(386), ra73.

+ Open protocol

+ Expand

Reverse Transcription and qPCR

Check if the same lab product or an alternative is used in the 5 most similar protocols

Annealing of oligodT primers to the RNAs is performed by incubating at 65°C for 5min 12 μL of RNA mixed with 0.5 μL of 100 μM oligodT(15) and 0.5 μL of 10mM dNTPs followed by an incubation 5min on ice. cDNAs synthesis was performed by adding 4 μL of 5X second strand buffer, 1 μL 0.1M dTT, 1 μL H₂O, 0.5 μL RNase inhibitor, and 0.5 μL superscriptIII reverse transcriptase (InVitrogen, 18080093) to the RNA solution and incubating at 50°C for one hour. The reverse transcription is stopped by adding 80 μL of H₂O to the reaction and incubating 10 min at 70°C. For qPCR, 5 μL of cDNAs were mixed with 12.5 μL SyberGreen (SIGMA, S9194), 7.22 μL H₂O, 0.14 μL each of Forward and Reverse primers (10 μM, see Table S8). The qPCR reaction was then performed on StepOnePlus System (applied biosystem) using the standard detection mode option.

Jullien J., Vodnala M., Pasque V., Oikawa M., Miyamoto K., Allen G., David S.A., Brochard V., Wang S., Bradshaw C., Koseki H., Sartorelli V., Beaujean N, & Gurdon J. (2017). Gene Resistance to Transcriptional Reprogramming following Nuclear Transfer Is Directly Mediated by Multiple Chromatin-Repressive Pathways. Molecular Cell, 65(5), 873-884.e8.

+ Open protocol

+ Expand

Validating miRNA and mRNA Expression

Check if the same lab product or an alternative is used in the 5 most similar protocols

To validate miR expression data, two step qRT-PCR, cDNA synthesis and real-time PCR amplification was performed. Minimum of 1–2 μg of total RNA was reverse transcribed by using Universal cDNA synthesis kit (Exiqon, USA) and specific miRs primers (Exiqon San Francisco, USA). Further miR PCR Kit (Exiqon, USA) amplification was performed in Roche RT-PCR machine. 5S-rRNA was used as an endogenous control for normalization.
To validate mRNA expression, qRT-PCR was performed in triplicate in ViiA7 real-time PCR machine (ABI, Whitefield, Bangalore, India) using Syber Green (Sigma, St. Louis, MO, USA) with specific primers for genes (Table 1). 18sRNA served as endogenous control for normalization. Relative quantification of miR and each gene was determined by calculating the Log RQ of each sample’s Ct value.

Trehanpati N., Sehgal R., Patra S., Vyas A., Vasudevan M., Khosla R., Khanam A., Kumar G., Maiwall R., Ramakrishna G., Kottilil S, & Sarin S.K. (2017). miRNA signatures can predict acute liver failure in hepatitis E infected pregnant females. Heliyon, 3(4), e00287.

+ Open protocol

+ Expand

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?

Revolutionizing how scientists
search and build protocols!