Home Protocols Appendix A for use with NEBNext Ultra II RNA First Strand Synthesis Module (E7771)

Appendix A for use with NEBNext Ultra II RNA First Strand Synthesis Module (E7771)

3.1. Fragmentation

Note: These recommendations have been optimized using Universal Human Reference Total RNA. Other types of RNA may require different fragmentation times.

Modified fragmentation times for longer RNA inserts.

Figure 3.1: Modified fragmentation times for longer RNA inserts. Bioanalyzer traces of RNA as shown in an RNA Pico Chip. mRNA isolated from Universal Human Reference RNA and fragmented with First Strand Synthesis Reaction Buffer and Random Primer Mix (2X) at 94°C for 5, 10 or 15 minutes, and purified using 2.2X volume of Agencourt RNAClean XP Beads or NEBNext RNA Sample Purification Beads. For libraries with RNA insert sizes larger than 300 bp, fragment RNA between 5–10 minutes.