Given that cytoplasmic miRNA processing by Drosha was discovered using alphaviral vectors,19,22 we hypothesized that alphaviral trans-amplifying RNA (taRNA) would be suitable for this purpose.24,25 Our recently developed second-generation taRNA vector system is bipartite, combining a nonreplicating ...
Levels of normal and mutant plectin mRNA were measured by semiquantitative real-time PCR (SQRT-PCR), amplifying complementary DNA (cDNA) from wild-type fibroblasts, transfected and non-transfected EBS-MD fibroblasts. A wild-type specific forward PCR primer was designed to discriminate between wild...
The GFP fragment (942 bp) was produced by amplifying the pEGFP-C1 plasmid using a polymerase chain reaction in the Proflex PCR system (ThermoFisher Scientific). The PCR product was purified using Monarch® Nucleic Acid Purification Kit (New England Biolabs Inc.). In addition, the 40-nt ...
We then measured HERV-K (HML-2) Env expression using different primers designed for nested PCR amplifying domains coding for either SU or TM in plasma before and after HIV-1 infection in subject OP-1830. Nested PCR was performed on plasma-isolated viral RNA. There was a strong signal at ...
The primer pairs for amplifying human HELB and GAPDH transcripts were ShHELB-2630; 5′ -GCTGGCCTGGAAGTAACTGT-3′ /AhHELB-2740; 5′ -AACTGTTTGCTCCTCGGACC-3′ and hGAPDH556/hGAPDH64211,14, respectively. Amplification was carried out initially for 1 min at 95 °C, ...
RT-PCR of lentiviral mRNA was performed using total RNA. The brain slices were lysed, and total RNA was extracted using the RNeasy Lipid Tissue kit (Qiagen, France) according to the manufacturer’s instructions. The RNA (1 μg) was denatured for 10 min at 68°C, and cDNA was generated...
Primer sequences for amplifying Myf5 and GAPDH are as described (Pan et al., 2011). Chromatin immunoprecipitation (ChIP) assays ChIP assays were performed with 10T1/2 cells overexpressing FLAG–Zic1 and Pax3–HA using the Fast ChIP method (Nelson et al., 2006) with some modifications. ...
PCRs targeted at amplifying DNA containing the MSY-1-binding site on the Glut 3 gene (-89 to +260 bp) from immunoprecipitated protein complexed to chromatin in the presence of an anti-MSY-1 antibody (MSY-1 Ab, lane 4) revealed a 354-bp region. Negative control consisted of no primary...
To the best of our knowledge, this is the first one-step pan-IAV real-time reverse-transcription PCR which has been validated by amplifying nine IAV However, standard Sanger sequencing being validated with virus isolation, and was used to detect IAV in poultry swabs under clinical diagnostic ...
(iii) amplifying the post-capture PCR library to produce one or more enriched HPV TrDNA; and (iv) optionally repeating steps (iii) and (iv); (d) adding at least one index to the one or more enriched HPV TrDNA; (e) performing multiplexed sequencing of the one or more enriched HPV ...