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Team:Stanford/Notebook/Lab Work/Week 3
From 2010.igem.org
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Spring: Brainstorming | Spring Meetings
Summer: Week 1 | Week 2 | Week 3 | Week 4 | Week 5 | Week 6 | Week 7 | Week 8 | Week 9 | Week 10 | Summaries
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7/5 Monday
Planning Meeting Notes
Agenda
- Design Updates
- Analog Comparator (Chris & Alex)
- Redundancy between the two proposals
- Degree of phosporylation
- Dynamic range and mechanisms
- Digital Comparator (Francisco & Karina)
- RSID; standardizing the design
- Introducing a PoPs signal output
- Characteristics between the two Comparators
- What makes the two different?
- Applications
- Vaginal Infections/Preterm Birth
- is it really a ratio?
- Other ideas: Targeting Cancer Cells, oscillators
- Administrative Details
- Ordering Stuff: Oligos ordering sheet, Digestion and Gel Extraction Kits?
- Stipend?
- Sponsorship Updates
- MDV Presentation
Goals For the Week
- Chris & Alex
- need to make 50x TAE
- complete plasmids as far can get w/o kinase and phosphotase
- order DNA sometime this week
- Francisco & Karina
- Run orginal and new designs by Christina
- design standard sRNA designs and place order by the end of the week
7/6 Tuesday
Greg
- During weekly meeting got up to speed with what the team’s been doing while I was away
- Used heat-shock transformation protocol for DH5alpha to transform with the following genetic material, left to culture overnight:
| Function | Part # | Distribution Location | Resistance |
|---|---|---|---|
| ribosome binding site | B0032 | P1 2I | Amp |
| forward double terminator | B0015 | P1 23L | Amp + Kan |
| bidirectional double terminator | B0014 | P2 24C | Amp + Kan |
| medium constitutive promoter | J23107 | P1 20A | Amp |
| strong constitutive promoter | J23100 | P1 18C | Amp |
| plasmid backbone | pSB1K3 | P1 5A | Kan |
7/8 Thursday
Christopher
Running a Gel of F2620, pBAD, and psb4A5:
- Make a 0.8% agarose gel (0.4 g of agarose in 50 ml of TAE)
- Heat in microwave until agarose is completely dissolved
- Add 10.0 ul of EtBr to mix (5000x for a 50 ml solution)
- let run for approximately 1 hr
- refer below for bands
4A5 E/P 3395 bp
I0500 E/S (on psb2K3) 1210 bp
F2620 E/S (on pSB1A2) 1061 bp
Gel Extract the Fragments Running a Gel of PCR Product:
- first need to PCR cleanup
- then run on 1.4% agarose gel (0.7 g of agarose in 50 ml of TAE)
- this is necessary because of the 800 bp size of the fragment
Miniprep Ligations
- F2620+E0240+psb1A2
- pBAD+E0240+psb1A2
Restriction Digests PCR product F2620
Laura
miniprepped Alex's/Chris' liquid cultures, using Promega kit
- 2 each of:
- psb1K3
- J23107
- B0032
- eluted in 100 uL H2O
| Nanodrop Data | |||
|---|---|---|---|
| tube ID | 260/280 | 260/230 | ng/uL |
| psB1K3-1 | 1.77 | 1.08 | 71.3 |
| psB1K3-2 | 1.73 | 1.01 | 77.8 |
| J23107-1 | 1.87 | 1.60 | 116.3 |
| J23107-2 | 1.90 | 1.74 | 142.0 |
| B0032-1 | 1.81 | 1.20 | 68.1 |
| B0032-2 | 1.83 | 1.32 | 72.5 |
7/9 Friday
Friday Recap Meeting
Weekly Presentation: Karina PPT
Analog Comparator update: Chris
MDV Presentation Updates & Planning
- July 16, 2010 11:00 am
- Separate Meeting for planning, Tuesday 4:00 pm, Green Atrium
- Grad Students: Ryan
Administrative Details
Stipend?
- Check on Axess for Billing statements
Sponsorship Updates
Christina’s announcements:
- Start talking about Medal Requirements
- Keep in mind the wiki!
- Digitized Cloning Scheme
Medal Requirements- Presentation next week how we’re addressing those
- Collaborative, Improving Standard, Human Practice
- Idea: improving promoter database on Parts Registry
- Need to get access to parts registry to edit e-mail Drew to e-mail them
- Gold Medal Meeting Meeting Wed Morning at 9:00 am? Send out the Doodle (Anusuya)
Next Weeks Leader: Alex
