User:Karmella Haynes/Notebook/ASU courses/2014/01/24: Difference between revisions

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{| {{table}}
| align="center" style="background:#f0f0f0;"|'''Colony'''
| align="center" style="background:#f0f0f0;"|'''1'''
| align="center" style="background:#f0f0f0;"|'''2'''
| align="center" style="background:#f0f0f0;"|'''3'''
| align="center" style="background:#f0f0f0;"|'''4'''
|-
| Conc.||30||227||243||45
|-
| 260/280||1.9||1.87||1.9||1.9
|-
|}
'''Digest plasmids with restriction enzymes and gel electrophoresis (1% agarose) to confirm the assembled construct size'''
{| {{table}}
| align="center" style="background:#f0f0f0;"|'''Plasmid DNA'''
| align="center" style="background:#f0f0f0;"|'''3.0 μL'''
|-
| EcoRI||1.0 μL
|-
| PstI||1.0 μL
|-
| 10x FastDigest buffer + green loading dye||1.5 μL
|-
| dH2O||8.5 μL
|-
| ||15.0 μL total
|-
| Incubate at 37°C for 10 minutes.||
|-
|}
[[Image:20140131 141956 (2).jpg|400px]]


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Revision as of 13:35, 3 February 2014

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01/24/14

  • Kozak plasmid minipreps

Kozak Plasmid Minipreps

  • For BME 100, spring 2014
  • Also for SynBERC Scholars/ Quanta "qOpenPCR" project

Day 1 - Tasks for Behzad:

  • Quick-transform Kozak miniprep into DH5α
  • Grow on 100μg/mL amp plates overnight. Include zero plasmid negative control

Day 2 - Tasks for Behzad

  • Pick 4 colonies, innoculate 4 cultures, 5 ml each
  • Grow overnight

Day 3 - Tasks for Behzad

  • Pellet the cells in the ancient table-top swing-bucket centrifuge
  • Very thoroughly (by vortexing and flicking and pipetting up and down) resuspend cells in appropriate amount of water
  • Proceed with miniprep
  • Measure DNA concentrations



Colony 1 2 3 4
Conc. 30 227 243 45
260/280 1.9 1.87 1.9 1.9

Digest plasmids with restriction enzymes and gel electrophoresis (1% agarose) to confirm the assembled construct size

Plasmid DNA 3.0 μL
EcoRI 1.0 μL
PstI 1.0 μL
10x FastDigest buffer + green loading dye 1.5 μL
dH2O 8.5 μL
15.0 μL total
Incubate at 37°C for 10 minutes.


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