Team:Stanford/Notebook/Lab Work/Week 3

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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


Contents

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