Biomolecular Breadboards:Protocols:3-PGA Buffer Prep: Difference between revisions
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==Preparation of 3-PGA Buffer for the Cell-Free Expression System, 14X== | ==Preparation of 3-PGA Buffer for the Cell-Free Expression System, 14X== | ||
{| border="1" cellpadding="2" | {| border="1" cellpadding="2" style="width: 600px" | ||
! scope="col | ! scope="col" | Component | ||
! scope="col | ! scope="col" | Stock | ||
! scope="col | ! scope="col" | Final Concentration in cell-free reaction | ||
! scope="col | ! scope="col" | 1-fold | ||
! scope="col | ! scope="col" | 4-fold | ||
! scope="col | ! scope="col" | 6-fold | ||
|- | |- | ||
| HEPES pH8 || 2M || 50mM || 150µl || 600µl || 900µl | | style="width: 75px;" | HEPES pH8 || style="width: 140px;" | 2M || style="width: 140px;" | 50mM || style="width: 75px;" | 150µl || style="width: 75px;" | 600µl || style="width: 75px;" | 900µl | ||
|- | |- | ||
| Water || || || 6µl || 24µl || 36µl | | Water || || || 6µl || 24µl || 36µl | ||
|- | |- | ||
| Homemade Nucleotides || 156mM: ATP & GTP | | Homemade Nucleotides || 156mM: ATP & GTP 94mM: CTP & UTP || 1.5mM: ATP & GTP 0.9mM: CTP & UTP || 57.8µl || 231.2µl || 346.8µl | ||
|- | |- | ||
| tRNA E.coli || 50mg/ml || 0.2mg/ml || 24µl || 96µl || 144µl | | tRNA E.coli || 50mg/ml || 0.2mg/ml || 24µl || 96µl || 144µl | ||
Revision as of 01:19, 6 June 2012
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Preparation of 3-PGA Buffer for the Cell-Free Expression System, 14X
| Component | Stock | Final Concentration in cell-free reaction | 1-fold | 4-fold | 6-fold |
|---|---|---|---|---|---|
| HEPES pH8 | 2M | 50mM | 150µl | 600µl | 900µl |
| Water | 6µl | 24µl | 36µl | ||
| Homemade Nucleotides | 156mM: ATP & GTP 94mM: CTP & UTP | 1.5mM: ATP & GTP 0.9mM: CTP & UTP | 57.8µl | 231.2µl | 346.8µl |
| tRNA E.coli | 50mg/ml | 0.2mg/ml | 24µl | 96µl | 144µl |
| CoA | 65mM | 0.26mM | 24µl | 96µl | 144µl |
| NAD | 175mM | 0.33mM | 11.5µl | 46µl | 69µl |
| cAMP | 650mM | 0.75mM | 7.1µl | 28.4µl | 42.6µl |
| Folinic Acid | 33.9mM | 0.068mM | 12µl | 48µl | 72µl |
| Spermidine | 1M | 1mM | 6µl | 24µl | 36µl |
| 3-PGA | 1.4M | 30mM | 128.6µl | 514.4µl | 771.6µl |
