IGEM:Harvard/2008/Lab Notebooks/DailyBook/Week0

Monday: June 16, 2008

Meeting Notes

Tic-Tac-Toe

Basic Idea

• Use a chemical signal to induce electric output
• Use E. coli as detector, Shewie as indicator
• Use E. coli to produce lactate, needed for e- production in Shewie

Logistics of Game

Person v. Person

• Example game:

1. Player 1 adds chemical
2. Computer tracks electric output and displays move on screen
3. Player 2 adds chemical
4. Computer tracks electric output and displays move on screen
5. Repeat

• One chemical detection needed

Person v. Computer

• Example game:

1. Player 1 adds chemical
2. Computer tracks electric output and displays move on screen
3. Computer chooses its move and lights up the well that it decides to move to
4. Bacteria respond in a certain way
5. Repeat

• One chemical detection and one light detection needed

Person v. Bacteria

• Most complex.
• Applying ideas from DNA paper to bacteria
• Create 8 strains of different antibiotic resistances
• More feasible in E. coli than in Shewie

Tuesday: June 17, 2008

• GFP/OD Measurement (results)
• Make glycerol stocks
• Miniprep vectors (protocol)
• Make competent Shewie
  • Chemically competent (protocol)
  • Electrocompetent (protocol)
• Pour CM plates
• Transform Shewie with pACYC Duet and GFP vectors

Name	Registry Name	Description	Origin	Size	Marker
E1	J23113	Low promoter GFP tester	pMB1	35	Kan
E2	J23150	Medium promoter GFP tester	pMB1	35	Kan
E3	J23151	High promoter GFP tester	pMB1	35	Kan
E4	pACYC duet	Low-Medium copy vector	p15a	4008	Cm

For list of all parts, see here.

Wednesday: June 18, 2008

• Cloning Strategy
  • Punch out from registry

Name	Registry Name	Description	Origin	Size	Marker
E5	pSB3K3	Low-Medium copy vector	p15a	2750	Kan
E6	BBa-E1010	RFP only	pMB1	681	Kan
E7	pSB1A2	GFP only	pMB1	2079	Amp
E8	BBa_J04450	RFP with LacI promoter	pMB1	1069	Amp
E9	BBa_J04430	GFP with LacI promoter	pMB1	1083	Amp
E10	BBa_I715038	T7 Polymerase with LacI promoter	pMB1	2878	Amp

For list of all parts, see here.

• transform into E. coli TOP10 cells

• Make chemically and electrocompetent Shewie MR-1
• Transform electrocompetent Shewie MR-1

Thursday: June 19, 2008

Restreaking electroporated E. coli plates

Some plates were overgrown (lawns/colonies too dense), so they were replated

• P4 pAYC-Duet (Cm) restreaked from lawn onto 1000x dilution chloramphenicol plate)
• P7 pSB1A2 one big and one small colony onto 2 ampicillin plates
• P8 BBa_J04450 one big and one small colony onto 2 ampicillin plates
• P9 BBa_J04430 one big and one small colony onto 2 ampicillin plates
• P10 BBa_I715038 one big and one small colony onto 2 ampicillin plates

Some plates didn't have any visible colonies