QuantSeq-Pool 3' mRNA-Seq Library Prep Kit - November 2021
Dear Customer
We are writing to notify you of an immediate update to our QuantSeq-Pool Sample-Barcoded 3' mRNA-Seq Library Prep Kit (Cat. No. 139.96). The update includes a product component change as well as protocol changes, which facilitate the following improvements:
Enhanced first strand synthesis buffer stability
Simplified protocol handling
The updated kits are provided with a revised User Guide (see Previous Version: 139UG272V0300) that is already available online.
We apologize for any inconvenience that this rapid product update may cause you. We are convinced however, that this will benefit improved long-term product quality and performance for you.
The updated QuantSeq-Pool kits are available from now on.
If you have any questions about these changes please contact support@lexogen.com. Please read on for further details.
Why was this upgrade required?
Our Quality Control Monitoring program detected a reduction in library pool yields after 1 year of storage. We have verified carefully that this yield decrease does not impact library data quality. The cause was traced to the First Strand cDNA Synthesis Mix (FS). To ensure robust stability of FS over the guaranteed shelf-life, we now provide an additional component, DTT that is combined with FS for First Strand cDNA Synthesis.
Product Component changes
FS composition was adjusted for increased stability.
Additional component DTT is provided.
Protocol changes
In addition to the component changes, our team verified that elevated temperature handling during first strand cDNA synthesis was not critical for library yield or data analysis quality. Therefore, an additional protocol update was also included.
In summary the specific protocol updates are:
Step 2: Reactions are cooled to, then held at 25 °C instead of 42 °C after denaturing the RNA / RT Primer mix.
Step 3: The mastermix is prepared with FS + DTT + Enzyme Mix 1 (E1). Prewarming of this mastermix at 42 °C is no longer required and was removed from the protocol.
ATTENTION! FS from kits that do not contain the DTT component (i.e., kits purchased before December 2021) cannot be combined with DTT using the updated QuantSeq-Pool protocol (139UG272V0300). In case of any questions please contact support@lexogen.com.
How does this update affect my library preps?
The final reaction volumes and composition of the first strand cDNA synthesis reaction do not change due to the separate addition of DTT.
The reduced holding temperature at Step 2 and omission of mastermix pre-warming has no negative impacts on library preparation or data quality.
There is no negative impact on data quality if libraries were already prepared with kits purchased prior to December 2021.