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NovaSeq2 PCR2 Redo

NovaSeq2 PCR2 Redo

Owned by Gregg Randolph
Last updated: Oct 08, 2020 by Muhammad Saqib
1 min read

5536 pcrs, 2 plates only get 10 columns

PCR2 Mastermix Preparation:

ul/rxn

Reagent

# of rxns

ul needed

ul/rxn

Reagent

# of rxns

ul needed

3

5X Phusion HF Buffer

5536

16800

0.45

10M dNTPs

5536

2520

0.3

Kapa HiFi HotStart DNA Pol

5536

1680*

0.5 ul

5 uM F and R FlowCell Primers

5536

2800

0.75

HPLC H2O

5536

4200

5

Total Volume

5536

28000

*Will need 7 tubes of Taq.

Distribute 145 ul to all wells of 2 plates. Use BenchSmart 96 well pipette to distribute 5 ul to 56 plates. Use 8 channel for the final two 10 column plates.

Use 96 channel to transfer 10ul PCR1 to appropriately labeled PCR2 redos.

qPCR Check:

qPCR gives a concentration of 12-16 nMoles/ul. Success! We will mail this to Psomagen on Monday.

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