while simple and often efficient, has two problems. First, the near-perfect overlap of the primers may promote self-annealing instead of pairing to the template. Second, the product is nicked at two sites and cannot be used as a template in subsequent PCR cycles, leading to poor yields. ...
Renal mRNA Expression of AQP1, AQP2, AQP3, and AQP4 as well as V2R During Kidney Development in Fetal Pigs Quantitative PCR (QPCR) revealed that renal AQP1, AQP2, AQP3, and AQP4 mRNA transcripts were detectable in 60 d gestation fetuses and the expression increased significantly a nz b...
PCR products were pooled as collections of 384 reactions from each plate, size-selected according to expected amplicon length, and used as a template for a second round of PCR to append plate-specific sample indexes and sequencing adapters. The second PCR was set using 10 μL of FailSafe ...
Quality assessment of total RNA based on A260/A280 ratio using the Nanodrop spectrometer (KAIAO, Beijing, China). Reverse transcription to complementary DNA (cDNA) using quality-tested RNA. cDNA template was obtained by reverse transcription using 5× PrimeScript RT Master Mix (Takara) reverse ...
The obtained cDNA served as a template for in vitro transcription with T7 RNA polymerase and biotin UTP to generate multiple copies of biotinylated cRNA. Each cRNA sample (1.5 μg) was hybridized overnight to a RatRef-12 BeadChip array (Illumina). The chips were then washed, dried, and ...
The raw gene expression data from the genes of interest were normalised using the geometric mean of the most stable candidate reference genes GAPDH, HPRT1, and RPL32. Depot-specific mRNA expression The present study investigated the AT depot related mRNA expression of inflammation-related genes in...
Template cDNA was synthesized from 2 μg total RNA using the RevertAid™ First Strand cDNA Synthesis Kit (Thermo Scientific) and random hexamer primers following manufacturer's instructions. Cycling conditions were 95 °C for 15min, followed by 45 cycles of 95 °C for 15 s, 56 °C for ...
The study was funded by Medical University of Lodz (project no. 503/5-064-01/503-1) and The National Centre of Science (grant nos. 2017/25/N/NZ5/02064 and 2017/27/B/NZ5/02011). Disclosure The authors report no conflicts of interest in this work. References 1...
Total RNA (1 mg) was subjected to two rounds of hybridization to oligo(dT) beads (Dynal, Carlsbad, CA, USA). The resulting mRNA was then used as a template for cDNA synthesis. The mRNA was randomly fragmented to between 200 and 700 bp by focused acoustic shearing (Covaris, Woburn, MA...
(cRBPs), or serve as templates for translation. The majority of intronic circRNAs (ciRNAs) and exon-intron circRNAs (EIciRNAs) are retained in the nucleus [16]. The circular form of the introns of C9ORF72 transcripts can be exported to the cytoplasm and serve as the translation template ...