Matt:LabNotes/2014-1-30
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Block Shifted
- Block Shifted FISSEQ_2ndRCAprimerDye with RT primer oligonucleotide to see if they are hybridizing to the same locations on primary rolonies
Procedure
- Stripped with 80% formamide for 10min at 50C
- Hybridize RT primer 1uM (2ul 100uM Dye + 60ul 100% formamide + 20ul 20X SSC + 118ul H2O)
- Wash with 2X SSC twice
- Hybridize Shifted FISSEQ 2uM (2ul 200uM Dye + 60ul 100% formamide + 20ul 20X SSC + 118ul H2O)
- Wash with 2X SSC twice
- In the future, the blocking oligonucleotide should be atleast one order of magnitude greater in concentration
Results
24hrs | Blocked |
289 | 107 |
116 | 68 |
359 | 366 |
21370 | 3213 |