Digest and gel verification (hEF1a:LacI, TRE:mKate, pLac:mKate, hEF1a:CD79A-XTCSX-gal4VP16)
enzymes:
| Ladder | pEXPR 1-Hef1a:LacI-2 | pEXPR 1-TRE mKate-2 | pEXPR 1-pLac mKate-2 | hef1a:cd79A-3T3-gal4 | hef1a:cd79A-XTCSX-gal4 |
|---|---|---|---|---|---|
| ApaI | SalI | ApaI | BglII, XbaI | BglII, MluI | |
| 9 | 10 | 11 | 12 | ||
 |  |  |  |  |  |
| Ladder, TRE:mKate, pLac:mKate, hEF1a:LacI, 3T3, 3T9, 3T15, 6T6, 6T9, 6T12 | Ladder, 9T3, 9T6, 9T9, 9T12, 9T12, 9T15, 12T6, 12T9, 12T12, 15T3, 15T9, 15T15 | Notes: |
 |  | The LRed constructs (1st 3 lanes after the ladder, left gel) seem to be in the wrong order. The band pattern in lane 1may be the failure mode of TRE:mKate, but there is an extra band at around 10kb which doesn't make sense, also there seems to be some undigested plasmid as well. The band pattern in lane 2 is probably the correct hEF1a:LacI, even though it says it's pLac:mKate. The band pattern in lane 3 is probably the correct pLac:mKate, even though it says it's hEF1a:LacI |
Sent For Sequencing:
| Sample1 | Sample2 | Sample3 | Sample4 | Sample5 | Sample6 | Sample7 | Sample8 | Sample9 | Sample10 | Sample11 | Sample12 | Sample13 | Sample14 | Sample15 |
|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
| S1-9T3 | S2-9T3 | S4-9T3 | S1-9T6 | S2-9T6 | S4-9T6 | S1-9T9 | S2-9T9 | S4-9T9 | S1-9T12 | S2-9T12 | S4-9T12 | S1-9T15 | S2-9T15 | S4-9T15 |
Gel verification of hEF1a:CD79B-pTet:mRFP-Gal4VP16 SDM
we got no bands on the gel. however, when we talked to nicholas (who has experience with SDM), he said that bands may or may not appear on a gel depending on how much amplification happened. So we're going to run the SDM again and transform the PCR product directly without running it on a gel.