LIMS verification - [Normalize RNA finished libraries on Hamilton and make dilutions in LIMS]

Work Flow: RNA v4

Protocol: Normalization of RNA samples for sequencing

Step: Normalization of RNA samples for sequencing

Description:

Describe the reason for the verification and other motivations

Finished libraries from the RNA prep at CG usually need dilution as we aim to sequence libraries with a concentration of 2.5 nM (around 0.5 ng/ul). This is calculated in a separate Excel sheet and then we normalize manually, which takes extra time and comes with human error and sample mixup risks compared to if LIMS did the calculations directly and we could normalize on Hamilton. Therefore, it would be a great enhancement for the method if the step Normalization of RNA samples for sequencing had these features.

Actions:

Describe/list actions to perform in order to complete the verification

[ ] An EPP called "Calculate volumes for dilution RNA" or similar where the user fills in process UDF "Final Concentration (ng/ul)", and the script would calculate and generate the volumes needed for dilution for the sample udfs "Sample Volume (ul)" and "Volume Buffer (ul)".
- If the concentration of a sample is <=1.5 ng/ul no dilution is needed and you can take 11 ul (the whole volume) of sample and 0 ul buffer.
- The total maximum volume (sample + buffer volume) should be 100 ul
- The minimum sample volume is 1 ul.
[ ] "Output container barcode" udf for the step Normalization of RNA samples for sequencing
[ ] "Output container barcode" udf for step A-tailing and Adapter ligation (RNA) to put on the "Finished Libraries" Biorad HSP plate
[ ] "Supporting procedure document" udf with "Hamilton Normalisation and pooling" and "NA" as alternatives in step Normalization of RNA samples for sequencing
[ ] "Hamilton instrument" udf with option "Linda" and "NA" in step Normalization of RNA samples for sequencing
[ ] Placement holder for the Hamilton norm file in step Normalization of RNA samples for sequencing
[ ] Epp "Generate Hamilton norm file" or similar in step Normalization of RNA samples for sequencing

Testing setup:

If doing more extensive testing, the setup and specification and results can be documented in the table below.

Protocol	Step	Test specification	Comments/Results	P/F	Sign
Normalization of RNA samples for sequencing	Normalization of RNA samples for sequencing	TBA	Comments or results from testing	P (Pass) or F (Fail) based on the criteria in the Test specification	Date and Sign

Project resources:

Don't forget to write where you document screenshots etc, if you keep them in a folder on Google Drive for example.

Conclusions:

Summarize conclusions here.

Sign off

Head of Unit Lab	Date

Implementation

What is needed to implement changes	Item	Date signature
Information to staff
Documentation
Other
Other

[ ] Verification completed and implemented

Clinical-Genomics / cg_lims