Improm 2 reverse transcription set

Manufactured by Promega

The Improm-II Reverse Transcription Set is a laboratory reagent used for the conversion of RNA into complementary DNA (cDNA) through the process of reverse transcription. The set includes the necessary components, such as a reverse transcriptase enzyme, required buffers, and other essential reagents to facilitate this fundamental step in molecular biology workflows.

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Lab products found in correlation

Sybr green primers, by Merck Group (1 mentions) Rneasy mini kit, by Qiagen (1 mentions) Stepone machine, by Thermo Fisher Scientific (1 mentions) Rt sybr green rox fast mastermix, by Qiagen (1 mentions) Nucleospin rna plus kit, by Macherey-Nagel (1 mentions)

2 protocols using improm 2 reverse transcription set

Quantifying Sema6C Expression by qRT-PCR

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The RNeasy Mini kit (Qiagen) was used to get total mRNA extracts. In a final volume of 20 μL, 1 μg of RNA was retrotranscribed using the Improm-II Reverse Transcription Set (Promega), according to the manufacturer’s protocol. Real-time PCR was used to evaluate gene expression using specific SYBR Green primers provided by Sigma-Aldrich Company (see table below). According to the formula below, the fold change was calculated: Fold increase = 2^-(CT of target gene − CT of the house-keeping gene). Primers applied were as follows:

h-Sema6C-S	5′-CTTCGGCTCAACTGCTCTGT
h-Sema6C-AS	5′-AACCCACGCTCAATCTCATC
h-GAPDH-S	5′-TTGTTGCCATCAATGACCC
h-GAPDH-AS	5′-CCTCCCGTTCTCAGCCTTG

Fard D., Testa E., Panzeri V., Rizzolio S., Bianchetti G., Napolitano V., Masciarelli S., Fazi F., Maulucci G., Scicchitano B.M., Sette C., Viscomi M.T, & Tamagnone L. (2023). SEMA6C: a novel adhesion-independent FAK and YAP activator, required for cancer cell viability and growth. Cellular and Molecular Life Sciences, 80(4), 111.

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Real-Time PCR Gene Expression Analysis

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Total RNA was extracted using a NucleoSpin-RNA Plus kit (Macherey-Nagel, #740984.50) according to manufacturer's instructions then converted to cDNA using the ImProm-II Reverse Transcription Set (Promega, #A3802). Two hundred nanograms of cDNA per reaction was used for real-time PCR on a StepOne machine (Applied Biosystems), with respective primers (shown below) and the RT² SYBR Green ROX FAST Mastermix (Qiagen, #330623). Amplification was performed at 95°C for 10 min, then 40 cycles of 15 s at 95°C and 1 min at 60°C. Relative mRNA quantifications were calculated using the established ΔΔCT method [58] with GADPH as housekeeping control. Results shown represent 2 -ΔΔCT values where one control sample per plate is arbitrarily equal to 1 (Table 1).

Lin H., Ji D.K., Lucherelli M.A., Reina G., Ippolito S., Samorì P, & Bianco A. (2020). Comparative Effects of Graphene and Molybdenum Disulfide on Human Macrophage Toxicity. Small (Weinheim an der Bergstrasse, Germany), 16(35).

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