Difference: ProceduresCrushSoakPolyacrylamideGelElution (1 vs. 3)
Revision 32012-06-06 - AlvaroRodriguez
ELUTION of OLIGOS FROM PAGE GEL - Crush SoakEluting DNA/RNA from PAGE or denaturing PAGEMaterials | ||||||||
| Changed: | ||||||||
| < < | Crush Soak Buffer (CSB) - 200mM NaCl , 10 mM tris-HCl (pH 7.5), 1mM EDTA (pH 8) | |||||||
| > > | Crush Soak Buffer (CSB) - 200mM NaCl , 10 mM tris-HCl (pH 7.5), 1mM EDTA (pH 8) using MilliQ water/NanoPure water. | |||||||
Procedure (Work in Progress)Elution * 1 Excise Band (try doing it with the least polyacrylamide from gel need to get whole band) with a clean razor. | ||||||||
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| < < | * 2 Transfer the excised band into an 1.7 microliter eppi tube and crush with a sterile pipet tip. | |||||||
| > > | * 2 Transfer the excised band into an 1.7 microliter eppi tube and crush into big chunks with a sterile pipet tip. | |||||||
| * 3 Add 2 volumes of CSB for about every 1 volume of polyacrylamide. * 4 Incubate overnight at 37 C and by shaking it. (Speeds the process) * After the incubation, centrifuge the sample at 15000 rpm for 2 minutes at 4 C. * Trasfer supernatant(has the DNA/RNA) carefully without aspirating any of the polyacrylamide to a new eppi tube. * 2X Add 2 volumes of CBS, repeat centrifugation step, and transfer the supernatant. * Ethanol Precipitate the DNA from the supernatant. | ||||||||
Revision 22012-04-13 - AlvaroRodriguez
ELUTION of OLIGOS FROM PAGE GEL - Crush SoakEluting DNA/RNA from PAGE or denaturing PAGE | ||||||||
| Changed: | ||||||||
| < < | Materials | |||||||
| > > | Materials | |||||||
| Crush Soak Buffer (CSB) - 200mM NaCl , 10 mM tris-HCl (pH 7.5), 1mM EDTA (pH 8) | ||||||||
| Changed: | ||||||||
| < < | Procedure | |||||||
| > > | Procedure (Work in Progress) | |||||||
| Elution * 1 Excise Band (try doing it with the least polyacrylamide from gel need to get whole band) with a clean razor. * 2 Transfer the excised band into an 1.7 microliter eppi tube and crush with a sterile pipet tip. * 3 Add 2 volumes of CSB for about every 1 volume of polyacrylamide. * 4 Incubate overnight at 37 C and by shaking it. (Speeds the process) * After the incubation, centrifuge the sample at 15000 rpm for 2 minutes at 4 C. * Trasfer supernatant(has the DNA/RNA) carefully without aspirating any of the polyacrylamide to a new eppi tube. * 2X Add 2 volumes of CBS, repeat centrifugation step, and transfer the supernatant. * Ethanol Precipitate the DNA from the supernatant. | ||||||||
| Deleted: | ||||||||
| < < | ||||||||
Revision 12012-04-12 - AlvaroRodriguez
ELUTION of OLIGOS FROM PAGE GEL - Crush SoakEluting DNA/RNA from PAGE or denaturing PAGE Materials Crush Soak Buffer (CSB) - 200mM NaCl , 10 mM tris-HCl (pH 7.5), 1mM EDTA (pH 8) Procedure Elution * 1 Excise Band (try doing it with the least polyacrylamide from gel need to get whole band) with a clean razor. * 2 Transfer the excised band into an 1.7 microliter eppi tube and crush with a sterile pipet tip. * 3 Add 2 volumes of CSB for about every 1 volume of polyacrylamide. * 4 Incubate overnight at 37 C and by shaking it. (Speeds the process) * After the incubation, centrifuge the sample at 15000 rpm for 2 minutes at 4 C. * Trasfer supernatant(has the DNA/RNA) carefully without aspirating any of the polyacrylamide to a new eppi tube. * 2X Add 2 volumes of CBS, repeat centrifugation step, and transfer the supernatant. * Ethanol Precipitate the DNA from the supernatant. |
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