09 320 Lab _5 restriction digest friday

09 320 Lab _5 restriction digest friday - Lab #5 Plasmid...

Info iconThis preview shows pages 1–5. Sign up to view the full content.

View Full Document Right Arrow Icon
Lab #5 Plasmid Isolation & Restriction Digest Section 1332 7 Friday 1:00pm-4:00pm ZHS 359 Zheng Zhang Office hours: T/Th 11:00am-12:00 zhengz@usc.edu
Background image of page 1

Info iconThis preview has intentionally blurred sections. Sign up to view the full version.

View Full DocumentRight Arrow Icon
Objectives: to isolate pGEM and pGEM- luc from bacterial cells to perform restriction digest Cultured overnight Centrifuged to pellet cells. Stored at 4 o C. single colony single colony Last Week:
Background image of page 2
DNA Isolation by Alkaline Lysis Goal to free the plasmid DNA from the cell, leaving behind the E. coli chromosomal DNA, cell wall debris, proteins and other cell components. Method: differential precipitation Long strands of E. coli DNA, entangled in the remnants of lysed cells, are precipitated. Small circular plasmid DNA stay in solution Three basic steps: 1. growth of bacteria to amplify the plasmid 2. harvesting and lysis of the bacteria 3. purification of the plasmid DNA
Background image of page 3

Info iconThis preview has intentionally blurred sections. Sign up to view the full version.

View Full DocumentRight Arrow Icon
E. coli chromosome and plasmid DNA E coli chromosome Plasmid DNA Size large small Structure linear, broken circular Base (NaOH) reaction denatured intact Differential separation precipitated in pellet stay in solution DNA denaturation by strong base (NaOH) Berg, JM, Tymoczko, JL & Stryer, L. Biochemistry , W.H.Freeman and Company. New York, 6th Ed. (OH
Background image of page 4
Image of page 5
This is the end of the preview. Sign up to access the rest of the document.

Page1 / 15

09 320 Lab _5 restriction digest friday - Lab #5 Plasmid...

This preview shows document pages 1 - 5. Sign up to view the full document.

View Full Document Right Arrow Icon
Ask a homework question - tutors are online