Author: Malboeuf, Christine M.; Yang, Xiao; Charlebois, Patrick; Qu, James; Berlin, Aaron M.; Casali, Monica; Pesko, Kendra N.; Boutwell, Christian L.; DeVincenzo, John P.; Ebel, Gregory D.; Allen, Todd M.; Zody, Michael C.; Henn, Matthew R.; Levin, Joshua Z.
Title: Complete viral RNA genome sequencing of ultra-low copy samples by sequence-independent amplification Document date: 2012_9_8
ID: s76c5ebd_14
Snippet: For both Ovation RNA-Seq version 1 and 2 systems (NuGEN, San Carlos, CA), viral RNA was amplified into dsDNA per manufacturer's protocol. Briefly, RNA was reversed transcribed into cDNA using oligo-dT and random primers, dsDNA was generated and amplified using single-primer isothermal linear amplification (SPIA), as previously described (34) . The following modifications were performed. First, we used 1.4 volumes of AMPure RNA clean beads prior t.....
Document: For both Ovation RNA-Seq version 1 and 2 systems (NuGEN, San Carlos, CA), viral RNA was amplified into dsDNA per manufacturer's protocol. Briefly, RNA was reversed transcribed into cDNA using oligo-dT and random primers, dsDNA was generated and amplified using single-primer isothermal linear amplification (SPIA), as previously described (34) . The following modifications were performed. First, we used 1.4 volumes of AMPure RNA clean beads prior to SPIA amplification. Second, the final amplified products were purified using 0.8 volumes of AMPure XP beads (Beckman Coulter Genomics, Danvers, MA). Amplified products were eluted in 30 ml TE buffer (Life Technologies). Technical replicates were performed for HIV NL4-3 clone, HIV clinical samples and WNV clone dilutions samples.
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