What RNA quality is recommended for the SMARTer smRNA-Seq Kit for Illumina?

High integrity RNA carrying intact 3'-OH is recommended. Compromised RNA templates may require dephosphorylation of the 3'-end prior to library preparation to enable the initial polyadenylation step of the kit workflow, requiring the presence of intact 3'-OH. smRNA-Seq [...]

What are the recommended RNA input quantities for the SMARTer® smRNA-Seq Kit for Illumina?

Inputs in the range of 1 ng to 2 µg of total or enriched small RNA are recommended.smRNA-Seq [...]

What type of Covaris machine and shearing parameters should I use?

Covaris shearing parameters for the SMART-Seq cDNA , were optimized using a Covaris S220 Focused-ultrasonicator to obtain size distribution of 100–500 bp with a peak of approximately 200 b. Use 130-?l Covaris tube, containing 75 ?l SMART-Seq cDNA in the Elution Buffer, set-up the following parameters: Peak Power – 175, Duty-10%, Burst Cycle – 200, Time(min)-5 min, Mode-Frequency Sweeping [...]

How do I convert double-stranded SMART-Seq mRNA cDNA into the sequencing library?

SMART-Seq mRNA cDNA can be converetd into the Illumina sequencing library using Takara Bio USA SMART Seq Library Prep Kit, Covaris shearing followed by library construction using the ThruPLEX DNA-Seq Kit, Illumina Nextera® XT DNA Sample Preparation Kit (using as input 100-300pg SMART Seq cDNA and following the standard Nextera XT protocol)Ultra-low input NGS-RNA [...]

What is the expected Gene Body Covergae of the SMART-Seq mRNA cDNA?

A combination of the SMART-Seq mRNA cDNA synthesis and Nextera XT library construction generated a slight 5' bias, as shown in the Technical Note Improving the sensitivity of ultra-low input mRNA-seq - Gene body coverage is good for all three library preparation methodsUltra-low input NGS-RNA [...]