The authors used gBlocks® Gene Fragments to generate a standard curve for their qPCR analysis.
PrimeTime qPCR Assays (primers and probes) for TRAAK, TASK-1, TASK-2, TALK-1, TRESK-2, and GAPDH were used in this study. Experiments were repeated a minimum of three times. The 2ΔΔCt method was used to calculate fold changes in gene expression.
Real-time PCR was performed using SYBR Green intercalating dye and primers. The primers were designed using the IDT PrimerQuest Primer Design Tool and synthesized by IDT. Relative gene expression (target gene expression normalized to the expression of the endogenous control gene) was calculated using the comparative Ct method (2−ΔΔCt).
The authors used gBlocks® Gene Fragments to generate a standard curve for their qPCR analysis.
Target-capture using xGen Lockdown Probes was combined with throughput RNA sequencing to provide high resolution detection of known gene fusions. The targeted gene fusions were known to be associated with several childhood sarcomas.
xGen Lockdown Probes were used to target an entire coding region, exon-intron boundaries, and segments of DNA containing known pathogenic variants in BRCA1 and BRCA2 genes.
Changes in short RNA abundance were tested for their possible influence on mRNA levels of 10 relevant genes. RT-qPCR expression analysis was performed on the 10 genes that displayed clear changes in short RNA read numbers in both sense and antisense orientation and their neighboring genes. cDNA was amplified using PrimeTime qPCR Assays (primers and 5' hydrolysis probes). Expression values represented the ΔCt values compared to β-actin.
The expression of 9 genes [Actn2, Actn3, Casq1, Drd2, Lepr, Mc4r, Mstn, Papss2, and Glut4 (a.k.a. Slc2a4)] with putative roles in the regulation of voluntary physical activity was evaluated via RT-qPCR of skeletal muscle and brain RNA from inherently high- and low-active mice. cDNA was amplified using PrimeTime Predesigned qPCR Assays, which included ZEN Double-Quenched Probes, with all reactions run in duplicate. Expression was normalized to an endogenous control (18S ribosomal RNA (IDT assay: RN18S) using methods described by Pfaffl.
Immunoprecipitated DNA was analyzed by real-time PCR using IDT PrimeTime qPCR Assays containing a ZEN Double-Quenched Probes and primers.
gBlocks Gene Fragments were used to generate mutant autonomously replicating sequences, or ARSs, associated with replication origins in yeast chromosomal DNA.
A gBlocks Gene Fragment was used to create an sgRNA expression vector for use with the CRISPR/Cas9 system.
gBlocks Gene Fragments were used to create sgRNA expression plasmids.
An RT-qPCR assay for HCV quantification was performed with probe-based assays using IDT primers and ZEN double-quenched probes.The assay amplified the 5′ UTR of the HCV genome and provided reduced background and increased signal.
The authors explain how next generation sequencing has advanced mitochondrial genetics research. They note how IDT’s xGen Lockdown Probes can be used to accomplish solution phase capture of mtDNA.