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ul/rxn | Reagent | # of rxns | ul needed |
|---|---|---|---|
10 ul | KAPA SYBR FAST qPCR MM (2X) | 6070 | 600700 |
2 ul | Primer Premix (10X) | 6070 | 120140 |
4 ul | Ultra Pure Water | 6070 | 240280 |
16 ul | Total Volume | 6070 | 9601120 |
Add 4 ul of template, pool, or standards to each well:
| 1 | 2 | 3 | 4 | 5 | 6 | 7 | 8 | 9 | 10 | 11 | 12 | ||
|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
A | ALF_PLT5_TN_16S_Col3 | ALF_PLT5_TN_16S_Col9ALF_PLT5_TN_ITS_Col3 | ALF_PLT5_TN_ITS_Col9ALF_PLT12_TN_16S_Col3 | ALF_PLT12_TN_16SITS_Col9Col3 | ALF_PLT12PLT5_TN12_ITS16S_Col3POOLALF_PLT12_TN_ITS_Col9 | HPAU_PP |
| NTC | NTC | NTC | ||||
B | ALF_PLT5_TN_16S_Col3ALF_PLT5_TN_16S_Col9 | ALF_PLT5_TN_ITS_Col3 | ALF_PLT5_TN_ITS_Col9ALF_PLT12_TN_16S_Col3 | ALF_PLT12_TN_16SITS_Col9Col3 | ALF_PLT12PLT5_TN12_ITS16S_Col3POOLALF_PLT12_TN_ITS_Col9 | HPAU_PP |
| 0.0002 pM Std | 0.0002 pM Std | 0.0002 pM Std | ||||
C | ALF_PLT5_TN_16S_Col3 | ALF_PLT5_TN_16S_Col9ALF_PLT5_TN_ITS_Col3 | ALF_PLT5PLT12_TN_ITS16S_Col9Col3 | ALF_PLT12_TN_16SITS_Col3 | ALF_PLT12PLT5_TN12_16S_Col9 | ALF_PLT12_TN_ITS_Col3 | ALF_PLT12_TN_ITS_Col9POOL_10000 | RHIR_PP_LN2 |
| 0.002 pM Std | 0.002 pM Std | 0.002 pM Std | ||
D | ALF_PLT5_TN_16S_Col3 | ALF_PLT5_TN_16S_Col9ALF_PLT5_TN_ITS_Col3 | ALF_PLT5PLT12_TN_ITS16S_Col9Col3 | ALF_PLT12_TN_16SITS_Col3 | ALF_PLT12PLT5_TN12_16S_Col9 | ALF_PLT12_TN_ITS_Col3 | ALF_PLT12_TN_ITS_Col9POOL_10000 | RHIR_PP_LN2 |
| 0.02 pM Std | 0.02 pM Std | 0.02 pM Std | ||
E | ALF_PLT5_TN_16S_Col3 | ALF_PLT5_TN_16S_Col9ALF_PLT5_TN_ITS_Col3ALF_PLT5_TN_ITS_Col9 | ALF_PLT12_TN_16S_Col3 | ALF_PLT12_TN_16SITS_Col9Col3 | ALF_PLT12PLT5_TN12_ITS_Col3POOLALF | RHIR_PLT12_TN_ITS_Col9PP_LN3 |
| 0.2 pM Std | 0.2 pM Std | 0.2 pM Std | ||||
F | ALF_PLT5_TN_16S_Col3 | ALF_PLT5_TN_16S_Col9ALF_PLT5_TN_ITS_Col3 | ALF_PLT5_TN_ITS_Col9ALF_PLT12_TN_16S_Col3 | ALF_PLT12_TN_16SITS_Col9Col3 | ALF_PLT12PLT5_TN12_ITS_Col3POOLALF | RHIR_PLT12_TN_ITS_Col9PP_LN3 |
| 2 pM Std | 2 pM Std | 2 pM Std | ||||
G | ALF_PLT5_TN_16S_Col3 | ALF_PLT5_TN_16S_Col9ALF_PLT5_TN_ITS_Col3 | ALF_PLT5_TN_ITS_Col9ALF_PLT12_TN_16S_Col3 | ALF_PLT12_TN_16S_Col9ALF_PLT12_TN_ITS_Col3 | ALF_PLT12PLT5_TN12_ITS_POOL_Col910000 |
| 20 pM Std | 20 pM Std | 20 pM Std | |||||
H | ALF_PLT5_TN_16S_Col3 | ALF_PLT5_TN_16S_Col9ALF_PLT5_TN_ITS_Col3ALF_PLT5_TN_ITS_Col9 | ALF_PLT12_TN_16S_Col3 | ALF_PLT12_TN_16S_Col9ALF_PLT12_TN_ITS_Col3 | ALF_PLT12PLT5_TN12_ITS_POOL_Col910000 |
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Results:
Average results are below:
ALF_PLT5_TN_16S_Col3:
ALF_PLT5_TN_16SITS_Col9Col3:
ALF_PLT5PLT12_TN_ITS16S_Col3:
ALF_PLT5PLT12_TN_ITS_Col9Col3:
ALF_PLT12PLT5_TN12_16S_Col3POOL:
ALF_PLT12PLT5_TN12_16S_POOL_Col910000:
ALF_PLT12PLT5_TN12_ITS_Col3POOL:
ALF_PLT12PLT5_TN12_ITS_Col9_POOL_10000:
HPAU_PP:
RHIR_PP_LN2:
RHIR_PP_LN3:
iSeq Run 16S POOL ONLY:
Dilute to 1 nM based off qPCR results. qPCR results are in pM, but 1:1000 dilution used. The results are effectively in nM for pool.
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