Daniel:Notebook/PosSequencing/2017-5-22
Jump to navigation
Jump to search
Single Stranded Probe Binding Test (Started Friday May 19)
Protocol (Continued from Saturday May 20)
- Dye hybridization
- Mix 20 uL ssDNA (lambda digested) and 20 uL dsDNA (original, non-digested template)
- Add 1 uL appropriate (10 uM) dye, A488-dc6 for sample 1 and Cy5-dc6 for sample 3
- Incubate in the dark (RT) for 15 minutes
- Magnet pulldown, remove supernatant and add 40 uL 2X SSC buffer
- Add 15 uL to a slide for analysis
- Seal coverslip with nail polish and wait approximately 30 minutes to dry
- Imaging Parameters
- Confocal Microscope
- 488 Laser-3.00%
- PMT Trans-Gain 225V
- PMT1-Range:500-550-Gain 550V
- PMT3-Range:750-800-Gain 600V
- 642 Laser-8.00%
- PMT Trans-Gain 300V
- PMT1-Range:500-550-Gain 6000V
- PMT3-Range:750-800-Gain 550V
Results
Raw Images
- 20170522-LambdaExonucleaseDigestion.lif Sample1-Field1-SNAP ch00.jpg
Sample 1 Field 1
- 20170522-LambdaExonucleaseDigestion.lif Sample1-Field2-SNAP ch00.jpg
Sample 1 Field 2
- 20170522-LambdaExonucleaseDigestion.lif Sample1-Field3-SNAP ch00.jpg
Sample 1 Field 3
- 20170522-LambdaExonucleaseDigestion.lif Sample1-Field4-Cy5-SNAP ch00.jpg
Sample 1 Field 4-Cy5
- 20170522-LambdaExonucleaseDigestion.lif Sample2-Field1-SNAP ch00.jpg
Sample 2 Field 1
- 20170522-LambdaExonucleaseDigestion.lif Sample2-Field2-SNAP ch00.jpg
Sample 2 Field 2
- 20170522-LambdaExonucleaseDigestion.lif Sample2-Field3-SNAP ch00.jpg
Sample 2 Field 3
- 20170522-LambdaExonucleaseDigestion.lif Sample2-Field3-Cy5-SNAP ch00.jpg
Sample 2 Field 3-A488