NoYES. QuantSeq FWD and REV libraries are prepared using the same protocol steps using different reagents and reaction volumes. The use of
When Globin or BC1 Block Blockers (RS-GBHs / RS-GBSs / RS-BC1B) or UMI Module (USS), is performed are added to the preps, the protocol remains unchanged and therefore the same script (QuantSeq FWD or REV) can be applied. This is because blockers are introduced by exchanging these module solutions with the standard RNA Removal Solution (RS) at step 5.
The same applies for the UMI Module (USS), or where the Second Strand Synthesis Mix 1 (SS1) at the respective steps (5 and 7). The transferred volume is the same at each step so only the supplied reagent tube needs to be exchanged – no protocol or volume changes are required. Therefore, the same automation protocol can be used for preparing all types of QuantSeq FWD and REV libraries, with or without Globin Block and / or UMIs.
Please note is exchanged at step 7 of the QuantSeq FWD protocol. The UMI module is not compatible with the QuantSeq REV kit.
NOTE: QuantSeq Flex libraries may require different automated protocol modifications depending on the use of custom primers at either or both of the first and second strand synthesis steps of library generation. QuantSeq Flex programming is available for the Biomek i7 and NGS Workstation B (Agilent Bravo). Please contact support@lexogen.com if you plan on automating QuantSeq Flex on any liquid handling platforms for more information.