Entry title
- PCR - validation of Chromatin sensor insert
PCR - validation of Chromatin sensor insert
- Designed primers to amplify three overlapping regions across the transgenes: Chromosensor1, Chromosensor 2
- chrsn1 f1/ chrsn1 r1; Validation region 1; sensor 1 (and 2)
- chrsn1 f2/ chrsn1 r2; Validation region 1; sensor 1 (and 2)
- Round 1: Control templates to test primer function
- U-2 OS genomic DNA (neg)
- Chromosensor 1 plasmid DNA
- Chromosensor 2 plasmid DNA
- Round 2: Genomic DNA prepped from candidate clonal lines
- To be done after chosing a single primer pair per validation region
- BD###, Chromosensor 1
- BD###, Chromosensor 1
- BD###, Chromosensor 2
- BD###, Chromosensor 2
ROUND 1 PCR: Neg, pos controls
Reagent
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Vol
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Mix* (x #)
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Expected: 1. CMV = 588
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Hover name 5 μL/lane; 1% agarose; Ladder
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genomic OR plasmid DNA |
1.0 / 0.5 |
7.0 / 4.5
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10 μM forward primer |
1.0 |
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10 μM reverse primer |
1.0 |
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2x GoTaq green |
10.0 |
70.0
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dH2O |
7.0 / 7.5 |
49.0 / 52.5
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20.0 μL |
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Note: *Prep one Mix per template
- For each template, aliquot 18.0 master mix into 6 tubes
Thermal cycler: Labnet - GOTAQ
- 95°C 3 min.
- 30x[95°C, 30 sec; 57°C 30 sec; 72°C 30 sec]
- 72°C 3 min.
- 4°C, ∞
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