If I have low cDNA concentration, how do I re-amplify it?

Protocol for re-amplifying the cDNA if concentration is too low by Qubit quantification

1. Create a cDNA master mix following the table below:

1. Pipet mix. 
2. Aliquot 45 uL of the master mix into a new PCR tube.
3. Add X uL of your cDNA template into the tube.
   1. NOTE: we recommend adding all your remaining cDNA into the reaction to make sure you get enough coverage
4. Mix well by pipetting. 
5. Briefly spin down the PCR tube.
6. Run the following PCR program (choose a cycle number most appropriate for your sample; e.g. if you had very little to no cDNA, run closer to 6 cycles):

1. Vortex the SPRI beads and add 30 µl to each of the tube (0.6X volume of the reaction mixture).
2. Vortex to mix for 10-15 seconds.
3. Incubate at room temperature for 5 minutes.
4. Briefly centrifuge the tubes and place the tubes on the magnetic rack. Once the solution is clear, carefully aspirate, and discard the supernatant.
5. Keeping the tubes on the magnetic stand, add 200 µl of 80% ethanol.
6. Wait 30 seconds and remove the supernatant.
7. Add 200 µl of 80% Ethanol.
8. Wait 30 seconds and remove the supernatant.
9. Briefly spin the tubes to collect the remaining ethanol at the bottom of the tubes.
10. Place the tubes back on the magnetic rack and remove the remaining ethanol.
11. Let the SPRI beads dry at room temperature until the beads appear matte (~1-2 minutes) but do not over dry.
12. Remove the tubes from the magnetic rack and add 10 µl of nuclease free water to each tube to elute. Pipette the beads to mix well and incubate at room temperature for 1 minute.
13. Place the tubes on a magnetic rack and incubate for 1 minute.
14. Transfer the supernatant to a new 0.2ml PCR tube or tubes.
15. Quantify the cDNA products using Qubit (1X dsDNA HS assay kit) and a Bioanalyzer or TapeStation following manufacturer’s guidelines (Bioanalyzer High Sensitivity DNA assay or TapeStation High Sensitivity D5000 assay).