calculate volume script in TGA workflow for normalisation

[EmiliaRegazzoni]

Lims Issue

For definitions of headings below see Basic Concepts.

Work Flow: TWIST v7 Protocol: Normalization of samples for sequencing (TWIST) Step: Normalization of samples for sequencing (TWIST)

Description: We would like a script that can calculate dilution volumes during normalisation of samples before sequencing. There are more details in the verification: https://github.com/Clinical-Genomics/Lab-projects/issues/519. The formula will be:

Target concentration (C2) in nM (OBS not ng/ul) = 4 nM Target volume (V2) = 30 ul Sample volume (V1) = ???????? Sample concentration (C1) in nM (OBS not ng/ul) = already calculated in the aggregation step (e.g. https://clinical-lims.scilifelab.se/clarity/work-complete/349113) Library Validation QC TWIST (post hyb) > Aggregate QC (Library Validation) TWIST Volume diluent (V3) = V2-V1

Where:

V1 = (V2*C2)/C1

and

V3 = V2-V1

Should be calculated by the script. For this to work, the nM concentration should replace the ng/uL concentration that is currently in the step. Furthermore we would like to remove the option to type in the final target concentration (in the example above and attached as a screenshot 1.5 ng/ul has been entered).

We would also like the script to warn the user if the volume required is too large or too small in the following way:

If the C1 is already <2, warn the user with: "Possible failed capture - Consider re-hyb - No dilution necessary", make V1 = 15 ul and V3 = 0

If the C1 <=6 but more than or equal to 2,"Dilution not required", make V1 = 15 ul and V3 = 0

If V1 is larger than than 15 ul, half the volumes once (as in use the target volume 15 instead of 30 and perform the calculations accordingly).

and lastly,

If V1 is less than or equal to 2 ul, double the volume once (as in use a target volume of 60 uL instead pf 30). If the V1 is STILL 2 ul or less, then warn the user but do not double the volume again.

Please contact Emilia if you have any questions.

[RasmusBurge-CG]

Great suggestion! 👏

[idalindegaard]

@EmiliaRegazzoni Is this still needed?

[EmiliaRegazzoni]

The though was that the dilution volumes would be available in lims if we do this - that would be nice - but on the other hand, it is nice to be able to dilute before the TS is done since we now have to finish as quickly as possible for sequencing... @linnealafleur @MarcelaGa @VasKyriak whaat do you think? this step is otherwise quite useless in lims