Blue:RNA-Seq Experiments:07282014: Difference between revisions
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* Yield for the RNA is too high (expect more of that seen for the 100-500 nuclei), therefore repeated PCR using 3 less cycles (18 total) | * Yield for the RNA is too high (expect more of that seen for the 100-500 nuclei), therefore repeated PCR using 3 less cycles (18 total) | ||
{| {{table}} border | |||
| align="center" style="background:#f0f0f0;"|'''Sample''' | |||
| align="center" style="background:#f0f0f0;"|'''Sample Concentration (ng/ul)''' | |||
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| NC||0.396 | |||
|- | |||
| RNA 1||11.5 | |||
|- | |||
| RNA 2||4.34 | |||
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Revision as of 17:02, 30 July 2014
SmartPlus on Mouse Nuclei
Obtained sorted nuclei from Aran:
- 100 nuclei sorted directly into 2ul clontech dilution buffer (4x)
- 500 nuclei sorted directly into 2ul clontech dilution buffer (4x)
- 5000 nuclei sorted into BSA buffer (2x)
- stored 100/500 sorts at -80C. Vacuum spun down to 1ul for each.
- RNA extraction of 5000 using Zymo Micro RNA Kit (including DNAse treatment). Eluted in 6ul and vacuum spun down to 2ul.
- Performed SmartPlus protocol as per C1 tube control protocol on 1ul of above samples (only modification is addition of randomer) using 21 cycles for PCR
- cDNA Yields:
Sample | Sample Concentration (ng/ul) |
NC | 0.999 |
RNA 1 | 72.1 |
RNA 2 | 40.0 |
100 nuc 1 | 7.93 |
100 nuc 2 | 8.75 |
500 nuc 1 | 5.20 |
500 nuc 2 | 6.25 |
- Yield for the RNA is too high (expect more of that seen for the 100-500 nuclei), therefore repeated PCR using 3 less cycles (18 total)
Sample | Sample Concentration (ng/ul) |
NC | 0.396 |
RNA 1 | 11.5 |
RNA 2 | 4.34 |