Difference between revisions of "Team:WashU StLouis/Notebook"
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| − | <h1>Notebook</h1> | + | <div class = "title2"></div> |
| − | <h2>Reading about labwork is <i> almost </i> as fun as doing labwork</h2> | + | <div class="action"><img src="https://static.igem.org/mediawiki/2016/d/d9/T--WashU_StLouis--Action.png" style="width:100%;"></div> |
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| + | <h1 class="actionwords">Notebook</h1> | ||
| + | <h2 class="speechwords">Reading about labwork is <i> almost </i> as fun as doing labwork</h2> | ||
| + | |||
<div class = "calendar-wrapper"> | <div class = "calendar-wrapper"> | ||
Revision as of 05:09, 19 October 2016
MENU ▤
Notebook
Reading about labwork is almost as fun as doing labwork
| June | ||||||
|---|---|---|---|---|---|---|
| 1 | 2 | 3 | 4 | |||
| 5 | 6 | 7 | 8 | 9 | 10 | 11 |
| 12 | 13 | 14 | 15 | 16 | 17 | 18 |
| 19 | 20 | 21 | 22 | 23 | 24 | 25 |
| 26 | 27 | 28 | 29 | 30 | ||
| July | ||||||
|---|---|---|---|---|---|---|
| 1 | 2 | |||||
| 3 | 4 | 5 | 6 | 7 | 8 | 9 |
| 10 | 11 | 12 | 13 | 14 | 15 | 16 |
| 17 | 18 | 19 | 20 | 21 | 22 | 23 |
| 24 | 25 | 26 | 27 | 28 | 29 | 30 |
| 31 | ||||||
| August | ||||||
|---|---|---|---|---|---|---|
| 1 | 2 | 3 | 4 | 5 | 6 | |
| 7 | 8 | 9 | 10 | 11 | 12 | 13 |
| 14 | 15 | 16 | 17 | 18 | 19 | 20 |
| 21 | 22 | 23 | 24 | 25 | 26 | 27 |
| 28 | 29 | 30 | 31 | |||
| September | ||||||
|---|---|---|---|---|---|---|
| 1 | 2 | 3 | ||||
| 4 | 5 | 6 | 7 | 8 | 9 | 10 |
| 11 | 12 | 13 | 14 | 15 | 16 | 17 |
| 18 | 19 | 20 | 21 | 22 | 23 | 24 |
| 25 | 26 | 27 | 28 | 29 | 30 | |
| October | ||||||
|---|---|---|---|---|---|---|
| 1 | ||||||
| 2 | 3 | 4 | 5 | 6 | 7 | 8 |
| 9 | 10 | 11 | 12 | 13 | 14 | 15 |
| 16 | 17 | 18 | 19 | 20 | 21 | 22 |
| 23 | 24 | 25 | 26 | 27 | 28 | 29 |
| 30 | 31 | |||||
1 / 30
No lab work done
June 1st
2 / 30
No lab work done
June 2nd
3 / 30
- First day in lab!
- Made 1L of LB Media
June 3rd
4 / 30
No lab work done
June 4th
5 / 30
No lab work done
June 5th
6 / 30
No lab work done
June 6th
7 / 30
- Performed gDNA Extraction on E. Coli MG1655 and Synechocystis 6803
June 7th
8 / 30
- Designed PCR primers for pgk, pck, fldA, petF, ATP Synthase 1, and ATP Synthase 2
June 8th
9 / 30
No lab work done
June 9th
10 / 30
- Primers Arrived
- Ran PCR on ATP Synthase 2 , pgk, fldA, petF, and pdCas9 plasmid
- Ran all PCR products on gel and recovered ATP Synthase 2, fldA, and petF
- Ran gradient PCR on pdCas9 plasmid
June 10th
11 / 30
No lab work done
June 11th
12 / 30
No lab work done
June 12th
13 / 30
Made and ran gel for pdCas9
- Ran PCR on ATP Synthase 1, pgk, and pck
- Performed gel extraction on yesterday’s ATP Synthase 2
- Ran pck, pgk, and yesterday’s gradient pdCas9 PCR products on gel and recovered all genes
- Performed gel extraction on pdCas9, fldA, petF
- Ran PCR on psc101 and psc102 plasmid
- Ran psc101 and psc102 PCR products on gel - were recovered
- Ran gradient PCR on ATP Synthase 1
- Made agar plates (forgot the antibiotic)
- Ran golden gate one pot assembly for fldA petF genes and the pdCas9 plasmid
June 13th
14 / 30
- Ran ATP Synthase 1 PCR products on gel - no bands seen
- Gel Extraction for pgk, pck, psc101, and psc102 plasmid
- Ran PCR on ATP Synthase 1 (3rd times the charm)
- Ran PCR products on gel - WE SAW BANDS! ATP SYNTHASE 1 RECOVERED
- Reran PCR on pgk, pck, psc101, and psc102 plasmid because we did not get high enough concentration during gel Extraction
- Made new agar plates (with the chloramphenicol)
- Transformed the petF, fldA plasmids into competent cells
- Plated colonies on agar plates and set in incubator
June 14th
15 / 30
- Checked for colonies (both genes were successful)
- Picked colonies and incubated overnight
- Ran yesterday’s PCR products on gel - bands were light but all were recovered
- Gel Extraction for pgk, pck, psc101, and psc102 plasmid
- Reran PCR on pgk, pck, psc101, and psc102 plasmid because we did not get high enough concentration during gel extraction
- Ran PCR products on gel - all were recovered
- Gel Extraction for pgk, pck, psc101, and psc102 plasmid
June 15th
16 / 30
- Miniprepped plasmids for fldA, petF
- Ran sequence PCR on both plasmids
- Ran PCR product on gel to confirm
- Clean and Concentrated PCR product
- Reran PCR on pgk, pck, psc101, and psc102 plasmid because we did not get high enough concentration during gel extraction (for the fourth time)
June 16th
17 / 30
- Prepared DNA for sequencing
- Delivered DNA to WashU Med School
- Added DPnI to psc101 and psc102 gel extract products, incubated for an hour, did Clean and Concentrate, measured conc
- Ran Golden Gate assembly on pck, pgk, and ATP Synthase
June 17th
18 / 30
- Electroporation with golden gate products
- Redid golden gate for pck, pgk, ATP synthase (did not run properly first time)
June 18th
19 / 30
- Electroporation with better golden gate products
- Picked colonies for worse golden gate for pck, pgk, ATP synthase
June 19th
20 / 30
- Miniprep worse ATP, pck, pgk golden gate
- Seq PCR worse ATP, pck, pgk golden gate
- Picked colonies for better golden gate for pck, pgk, ATP synthase
June 20th
21 / 30
- Made frozen stock of better golden gate pck, pgk, ATP synthase
- Miniprepped better golden gate pck, pgk, ATP synthase (low concentration)
June 21st
22 / 30
- Ran sequencing PCR for pgk, pck (4 colonies each)
- Gel Extraction for pgk, pck PCR product (no bands)
June 22nd
23 / 30
- Ran sequencing PCR for pck and pgk on a gradient
- Gel Extraction for pck PCR product was successful, not pgk
June 23rd
24 / 30
- Ran sequencing PCR for pgk on a different gradient
- Gel Extraction for pgk PCR product was not successful
June 24th
25 / 30
No lab work done
June 25th
26 / 30
- Picked four new colonies of pgk
- Started 4 new cultures from frozen stock
June 26th
27 / 30
- Made frozen stock of the 4 new colonies
- Miniprepped all 8 pgk cultures and measured concentrations
- Ran gradient PCR on all 8 pgk plasmids
- Prepped and sent ATP synthase and pck in for sequencing
June 27th
28 / 30
No lab work done
June 28th
29 / 30
- Mixed new pgk sequencing primers
June 29th
30 / 30
- Ran gradient PCR on all 8 pgk plasmids
June 30th
1 / 31
No lab work done
July 1st
2 / 31
No lab work done
July 2nd
3 / 31
No lab work done
July 3rd
4 / 31
4th of July!
July 4th
5 / 31
No lab work done
July 5th
6 / 31
- Sequencing results for ATP and pck were not good
- Reran seq PCR on ATP Synthase (bad results)
- Picked new ATP colonies
July 6th
7 / 31
- Miniprepped overnight cultures of ATP 5-8 and forgot to make frozen stock
- Ran PCR on ATP plasmids 5-8
- PCR for pgk with new primers with gradient
- Ran pgk PCR product on gels
- Ran golden gate on pgk PCR gel extract
July 7th
8 / 31
- Electroporation with golden gate pgk product
July 8th
9 / 31
- Picked 4 pgk products and inoculated them overnight
July 9th
10 / 31
- Made frozen stock of new 4 pgk cultures
- Miniprepped cultures
- Seq PCR on new cultures
- All 4 cultures showed correct bands
- Clean and Concentrate pgk PCR product
- Prepped pgk for sequencing
July 10th
11 / 31
- Delivered pgk for sequencing
July 11th
12 / 31
- New ATP synthase sequencing primers and pck regular primers came in
- Seq PCR of ATP synthase plasmids with new sequencing primers. No bands
- Picked 4 new ATP synthase colonies (again)
- Inoculated MG1655 from frozens stock to make more genomic DNA
July 12th
13 / 31
- Extracted genomic DNA from MG1655
- Miniprepped new ATP cultures
- Ran sequencing PCR on new ATP and got no bands at all. Will use newly transformed cells tomorrow
- Ran PCR with pck primers, got light bands
- Reran PCR with pck products
July 13th
14 / 31
- Ran pck on gel
- Gel extracted pck and combined gel extracts to improve concentration
- Golden gate ligation on pck
- Ran ATP sequencing PCR again with different backbone primers
July 14th
15 / 31
- Ran gel from yesterday’s ATP PCR. No bands
- Ran PCR for pfo
- Incubated pck and BioBrick cells
July 15th
16 / 31
- Miniprepped pck and biobrock genes (HSP)
July 16th
17 / 31
No lab work done
July 17th
18 / 31
- Ran ATP seq PCR gradient again. No bands. Done with ATP
- BioBrick digestion, ligation, transformation with HSP and GFP
- Ran PCR seq for pck and gel extract
July 18th
19 / 31
- Delivered pck sequence to med school
- Ligated backbone for BioBrick
- Extracted more genomic DNA from MG1655
- Gradient PCR on pfo
- Received 𝛥aceE cells from Yale
- Rehydrated and incubated 𝛥aceE cells
July 19th
20 / 31
- Ran PCR and gel extract for fldA backbone and petF backbone
- DPn1 the two backbones
- golden gate ligation on pfo gene and two backbones
- Streaked colony from 𝛥aceE plate onto LB/kan plate
July 20th
21 / 31
- Making media for competent cell protocol
- Made new LB with MgSO4 for chemical transformation
July 21st
22 / 31
- Digested BioBrick miniprep to check length
July 22nd
23 / 31
- Chemically competent cell protocol with 𝛥aceE
- Miniprepped pfo-petF and fldA-petF
- Seq PCR miniprepped pfo-petF and fldA=petF
- Remade LB with MgSO4 with correct concentration
July 23rd
24 / 31
No lab work done
July 24th
25 / 31
- Restarted comp cell protocol (3rd times the charm)
- Clean and Concentrate junction 2 for pfo-fldA
- Conducted HSP test in LB
July 25th
26 / 31
- Redid HSP test in minimal media M9
- Recultured BioBrick and DH10B control
July 26th
27 / 31
- Reset up BioBrick experiment but grew diluted cells with LB instead of M9, so we have to do it again
July 27th
28 / 31
- Redid HSP test in minimal media M9
July 28th
29 / 31
No lab work done
July 29th
30 / 31
No lab work done
July 30th
31 / 31
No lab work done
July 31st
1 / 31
- Chemical transformation of pfo-petF, pfo-fldA, petF, fldA into knockout strain, plated 80 and 240 ul, also control
- Part 1 of interlab study (LUDOX)
- Transforming 5 interlab plasmids
August 1st
2 / 31
- Picked and incubated electron donor colonies plated yesterday
- Picked and incubated interlab colonies plated yesterday (2 per construct)
August 2nd
3 / 31
- Made frozen stock of incubated colonies and interlab parts
- Redid BioBrick experiment at 37C
August 3rd
4 / 31
No lab work done
August 4th
5 / 31
No lab work done
August 5th
6 / 31
No lab work done
August 6th
7 / 31
- Started induction but had to stop bc no frozen stock of fldA-pfo or petF-fldA in DH10B
- Transformed fldA-pfo and petF-pfo into DH10B
August 7th
8 / 31
- Made frozen stock of 𝛥aceE
- Made 60% ethanol
- Incubated 8 constructs for biotin assay w/ methanol extraction (4 constructs and blank in DH10B and 𝛥aceE)
August 8th
9 / 31
- Methanol extraction assay (grew cells and froze pellets)
August 9th
10 / 31
- Assayed petF-aceE cells. Absorbance was too low to be meaningful
August 10th
11 / 31
- Ran preliminary sonication test
- Incubated large volume of DH10B
August 11th
12 / 31
- Ran large sonication test on DH10B to decide protocol
August 12th
13 / 31
- Incubated large volume of DH10B
August 13th
14 / 31
- More sonication tests
August 14th
15 / 31
- Made ATP standards from assay instructions
August 15th
16 / 31
- Made more LB
August 16th
17 / 31
- Inoculated pgk, pck, DH10B for assay tomorrow
August 17th
18 / 31
- ATP luminescence assay
- Inoculated 4 constructs and blank in 𝛥aceE cells for tomorrow
August 18th
19 / 31
- Nothing grew, couldn’t do induction
August 19th
20 / 31
- Inoculated DH10B constructs and control from frozen stock
August 20th
21 / 31
- DH10B electron donor induction, froze pellets, measured OD600
- Re-transformed constructs into 𝛥aceE cells
August 21st
22 / 31
- Sonicated and assayed pellets for biotin (1 minute, 40 amp, 1mL PBS)
- Reinoculated 𝛥aceE constructs in all antibiotic combinations to determine problem, in LB and plates
August 22nd
23 / 31
- Visited Monsanto
August 23rd
24 / 31
- Measured absorbance data from biotin in DH10B cells
- Did ATP Assay and got luminescence data
- Incubated all four 𝛥aceE electron donor constructs from frozen stock plates and original plates and incubated 𝛥aceE and DH10B
August 24th
25 / 31
- Nothing grew from yesterday's inoculations
- Interlab study
August 25th
26 / 31
- Remade competent cells
- Finished interlab study (FITC) and sent to iGEM HQ
August 26th
27 / 31
No lab work done
August 27th
28 / 31
No lab work done
August 28th
29 / 31
No lab work done
August 29th
30 / 31
No lab work done
August 30th
31 / 31
No lab work done
August 31st
1 / 30
No lab work done
September 1st
2 / 30
No lab work done
September 2nd
3 / 30
- Picked colonies from 4 new 𝛥aceE plates and incubated
September 3rd
4 / 30
- 𝛥aceE cell induction
September 4th
5 / 30
No lab work done
September 5th
6 / 30
No lab work done
September 6th
7 / 30
No lab work done
September 7th
8 / 30
No lab work done
September 8th
9 / 30
No lab work done
September 9th
10 / 30
No lab work done
September 10th
11 / 30
No lab work done
September 11th
12 / 30
No lab work done
September 12th
13 / 30
No lab work done
September 13th
14 / 30
No lab work done
September 14th
15 / 30
No lab work done
September 15th
16 / 30
- Incubated pck, pgk, DH10B from frozen stock
September 16th
17 / 30
- Ran ATP assay
- BioBrick digestion and ligation
- Sonicated 𝛥aceE pellets
- Ran 𝛥aceE cell absorbance scan
- Ran biotin assay on 𝛥aceE cells
September 17th
18 / 30
No lab work done
September 18th
19 / 30
No lab work done
September 19th
20 / 30
No lab work done
September 20th
21 / 30
No lab work done
September 21st
22 / 30
No lab work done
September 22nd
23 / 30
No lab work done
September 23rd
24 / 30
No lab work done
September 24th
25 / 30
No lab work done
September 25th
26 / 30
- Transformed ligated petF and pck BioBrick plasmids into DH10B (wrong antibiotic)
September 26th
27 / 30
No lab work done
September 27th
28 / 30
- Transformed ligated petF and pck BioBrick plasmids into DH10B (kan = right)
- Wrapped pck and pgk plasmids to be sent to Cardiff University iGEM
September 28th
29 / 30
- Small colonies grew on all plates
- Picked colonies and inoculated
- Sent plasmids to Cardiff University iGEM
September 29th
30 / 30
- Miniprep on pck and petF BioBrick
- Reusupended TetR-pTet gBlocks
September 30th
1 / 31
- Digestion/Ligation/Transformation of pSB1C3 and tetr-ptet BioBrick
October 1st
2 / 31
- TetR-pTet colonies grew, picked colonies
October 2nd
3 / 31
- Miniprepped TetR-pTet BioBrick
October 3rd
4 / 31
- digested BioBricks to check bands sizes
October 4th
5 / 31
No lab work done
October 5th
6 / 31
No lab work done
October 6th
7 / 31
No lab work done
October 7th
8 / 31
No lab work done
October 8th
9 / 31
No lab work done
October 9th
10 / 31
No lab work done
October 10th
11 / 31
No lab work done
October 11th
12 / 31
- Ran PCR on BioBrick plasmids
October 12th
13 / 31
- Ran PCR products on gel
- Only petF showed up
October 13th
14 / 31
- digestion of pck, TetR-pTet, and pSB1C3 with EcoRI
October 14th
15 / 31
- Digestion of pck, TetR-pTet, and pSB1C3 with PstI
- Ligated pck and TetR-pTet to pSB1C3
- Transformed into DH10B
October 15th
16 / 31
- Only TetR-pTet grew
- Ran colony PCR with 8 pck colonies and 8 TetR-pTet colonies
October 16th
17 / 31
- Ran colony PCR products on a gel but got no bands
- Miniprepped pTet1-8
- Ran PCR on pTet 1-8 plasmids to check if insert was present
- Ran PCR product on gel, got the band!
- Prepped petF, pck, and TetR-pTet to be sent to iGEM HQ
October 17th
18 / 31
- Last day in lab!
- Sent parts to iGEM!
October 18th
19 / 31
WIKIFREEZE
October 19th
20 / 31
Done in lab
October 20th
21 / 31
Prepping for Jamboree
October 21st
22 / 31
Prepping for Jamboree
October 22nd
23 / 31
Prepping for Jamboree
October 23rd
24 / 31
Prepping for Jamboree
October 24th
25 / 31
Prepping for Jamboree
October 25th
26 / 31
Prepping for Jamboree
October 26th
27 / 31
Fly to Boston for Jamboree
October 27th
28 / 31
GIANT JAMBOREE
October 28th
29 / 31
GIANT JAMBOREE
October 29th
30 / 31
PRESENTATION
October 30th
31 / 31
Fly back to St. Louis
October 31st
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