Document the dates you worked on your project.
What should this page have?
- Chronological notes of what your team is doing.
- Brief descriptions of daily important events.
- Pictures of your progress.
- Mention who participated in what task.
Inspiration
You can see what others teams have done to organize their notes:
Cancer Group
- PCR of 3xHA-TDGF1 construct. 12 cycles
- Gel of PCR Product → PCR was a failure. Band was 300bp. Should have been 750bp
- Second PCR of 3xHA-TDGF1 construct. 25 cycles instead of 12 → left overnight
Interlab Study
- Measured LUDOX and H2O for plate reader
- Transformation of positive and negative controls, test devices 1-3
Cancer Group
- Ran gel on PCR of yesterday's construct → Gel worked
- Gel purification of PCR Product → low concentration of DNA found during nanodrop
- PCR the construct using a revised method
- 2:54pm → ran the gel on the PCR product
Interlab Study
- Heat and "SB" streaking seems not to have worked
- No visible red colonies on plate
Cancer Group
- PCR Not working well → keep trying new methods with different annealing temperatures
- Tried 65 and 67 degrees C
Interlab Study
- FITC started
- Cutting re-inoculated
- Ordered new LUDOX
Other
- College of Arts and Sciences Pre College Summer Institute students stopped by lab
- Spoke to them about information on synthetic biology and iGEM
Cancer Group
- PCR worked → Test 5 → changed annealing temperature to 66 degrees C + increased the denaturing time
- Nanodropped DNA
Cancer Group
- Digestion of TDGF-1 construct and YFP352GAP vector with two restriction enzymes
- Will ligate later
|
TDGF-1 Construct (Phire PCR 128.2 ng/mL) |
YEP352GAP Vector (343.2 ng/mL) |
| Total Reaction Volume |
50 uL |
50 uL |
| Nuclease Free Water |
27.4 uL |
37.2 uL |
| 10x Cutsmart Buffer |
5 uL |
5uL |
| xho1 |
1 uL |
1 uL |
| Pvu II |
1 uL |
1 uL |
| DNA |
15.6 uL |
5.8 uL |
Cancer Group
- Running yesterday's digestion on gel
Week 2 (7/5-7/8)
7/5
Cancer Group
- Ran the PCR'ed construct on a gel with Phire Polymerase → It didn't work
Cancer group
- Ran the construct with PCR on gel → it worked
- Tasnia stabbed our gel
- Interlab study continues
Cancer group
- Nanodrops were pretty low (84-120 ng/nl)
- 11:46am PCR'ed the highest nanodrop result in tube #1 → PCR successful
- 2:33pm → attempt to digest with remaining non-PCR product → removed from incubator at 5:00pm
- Made a gel → ran PCR
Cancer group
- Ran PCR on the construct (5x) → gel did not work
- PCR'ed 84.2 ng/ml construct
- 5 replicates made using Phire
- Thermocycler Program
| Thermocycler Program |
| Step |
Temperature (Degrees C) |
Duration (seconds) |
| 1 |
98 |
30 |
| 2 |
98 |
20 |
| 3 |
66 |
5 |
| 4 |
72 |
30 |
| 5 |
72 |
60 |
| 6 |
4 |
Hold |
| After step 4, return to step 2. Repeat steps 2-4 thirty-five times before continuing to step 5 and on. |
| Water |
19.5 uL |
| Phire Polymerase |
25 uL |
| Template |
0.5 uL |
| Forward Primer |
2.5 uL |
| Reverse Primer |
2.5 uL |
- Restreaking of YEP352GAP transformed cells for miniprep
- Restreaking placed in incubator at 12am
Cancer group
- Made gel for electrophoresis of PCR construct → it didn't work
- Thermocycler Program: Same as PCR from 7/11
| Q5 Rx Buffer |
10 uL |
| 10nM dNTPs |
1 uL |
| 10 uM Forward Primer |
2.5 uL |
| 10 uM Reverse Primer |
2.5 uL |
| Template (Q5 PCR Pw.1 for 6/30/16) |
0.5 uL |
| Q5 Polymerase |
2.5 uL |
| Water |
33 uL |
- LB liquid culture (3ml with 3ul of ampicillin)
- Loaded 10ul Ladder DNA.
- Loaded 1ul dye + 5ul PCR product
- Ran gel at 115V
- Checked gel ladder → faint, no band for the PCR product
- Made new gel, loaded with the same amount of reagents as before → ran at 90V → No ladder band
- Ran it again at 115V → visible ladder, no PCR band
- Preparing a gel to run ladder and construct that previously showed a strong band → used this to diagnose a problem with gel setup. Ran TDGF1 152.9 Phire Child. Lane 1 is ladder, Lane 2 is that construct (10ul) with 2ul of dye
- Setup new PCR using the 6/30/16 PCR construct
- 98°C → 30sec
- 98°C → 20sec
- 66°C → 5sec
- 72°C → 30sec
- 72°C → 1min
- 4°C → hold
Cancer group
- Ran 5ul of PCR with ladder, cancer construct and vaccine construct → to diagnose problem with gel
- Miniprep of Dean vector (3ml LB culture)
- Purification of PCR product → nanodropped 16.4ug/ul
- Nanodrop of:
- Yellow 7/13 construct NEB purified → 16.4ng/ml
- Blue gel purification → 48.7ng/ul
- Pink PCR purification 7/13 → 102.2ng/ul
- 7/13 -RK Vector 1 → 48.2ng/ul
- 7/13 -RK Vector 2 → 96.3ng/ul
Vaccine Group
| Contents of well |
Amount (ul) |
| Ladder |
10 |
AD |
5 |
| AO |
5 |
| AM |
5 |
| AN |
5 |
| Gal-Sunil |
Added 1ul loading dye |
| Con-Sunil |
5 |
- Brian and Sunil's Gal promoters worked but both constructs failed
Biobrick ideas
- Smelly yeast → optimized from e. coli
- Twist on smelly e. coli on e. coli → regulated with quorum sensing
- OmpA/EnvZ recognize osmolarity
- Genetic Circuit
- Shine yellow light → Banana smell. Shine green light → wintergreen
- TetR operator system genetic circuit
- Have cell die at certain wavelength → killswitch
