Haynes Lab:Notebook/David Barclay Undergrad Training/Plasmid Transformation/2013/09/13

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Day 3- Minipreps

Liquid Culture Results

NOTE: Cultures and Streaks taken out after 11 hours (37 incubator) and left on lab bench for 11 hours.

hPCD

  • Cultures Innoculated 09/12/13
  • Cultures 1 and 2: Both show growth
  • Streak 1 and 2: Both show growth

Ubc

  • Cultures Innoculated 09/12/13
  • Cultures 1 and 2: Both show growth
  • Streak 1 and 2: Both show growth

Samples for Minipreps and Analysis

  • Sample 1= hPCD/V0120 colony 1; BBa_J176000; part= 186 bp; vector= 4968 bp
  • Sample 2= hPCD/V0120 colony 2; same
  • Sample 3= Ubc/V0120 colony 1; BBa_J176003; part= 368 bp; vector= 4968 bp
  • Sample 4= Ubc/V0120 colony 2; same
  • Sample 5= hPCD/V0120; BBa_J176000; part= 186 bp; vector= 4968 bp
  • Sample 6= Ubc/V0120; BBa_J176003; part= 368 bp; vector= 4968 bp

DNA Concentration Data

Plasmid OD260 OD260/280 ng/μL
1. hPCD 1 0.279 1.831 278.676
2. hPCD 2 0.242 1.834 241.754
3. Ubc 1 0.097 1.787 96.614
4. Ubc 2 0.146 1.789 145.548

Restriction Digest Table

  • Checked plasmid minipreps with EcoRI/PstI digests
Reagent Volume Expected:
1. hPCD = 186 bp, 4968 bp
2. Ubc = 368 bp, 4968 bp
Today's gel
15 μL/lane; 1% agarose; Ladder
DNA(plasmid) 3.0 μL
10X FD Buffer 1.5
EcoRI 1.0
PstI 1.0
dH2O 8.5
  15 μL --> 37°C/ 15 min.

Conclusion There were definite gel lines at ~4900 and faint lines at ~200 for hPCD (lanes 1,2,5) and ~ 350 for Ubc (3,4,6).
This signifies a working gel.

DNA Sequencing Samples

  • Submitted to DNASU on: '8/14/13'
  • Order Number: 8043
  1. Plasmid 1 (hPCD) - Forward P001
  2. Plasmid 1 (hPCD) - Reverse P002
  3. Plasmid 4 (Ubc) - Forward P001
  4. Plasmid 4 (Ubc) - Reverse P002


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