Metacaspase
Download
1 / 11

Metacaspase - PowerPoint PPT Presentation


  • 116 Views
  • Uploaded on

Metacaspase. Single or Multicopy gene in Emiliania huxleyi ?. Isolation of Genomic DNA from Emiliania huxleyi. CTAB Isolation and CsCl Gradient. Research Plan. Isolate Genomic DNA. Southern Blot Analysis. Digest Genomic DNA w/ Various Restriction Enzymes.

loader
I am the owner, or an agent authorized to act on behalf of the owner, of the copyrighted work described.
capcha
Download Presentation

PowerPoint Slideshow about ' Metacaspase' - palti


An Image/Link below is provided (as is) to download presentation

Download Policy: Content on the Website is provided to you AS IS for your information and personal use and may not be sold / licensed / shared on other websites without getting consent from its author.While downloading, if for some reason you are not able to download a presentation, the publisher may have deleted the file from their server.


- - - - - - - - - - - - - - - - - - - - - - - - - - E N D - - - - - - - - - - - - - - - - - - - - - - - - - -
Presentation Transcript
Metacaspase

Metacaspase

Single or Multicopy gene in Emiliania huxleyi?


Isolation of genomic dna from emiliania huxleyi

Isolation of Genomic DNA from Emiliania huxleyi

CTAB Isolation and CsCl Gradient


Research plan
Research Plan

Isolate Genomic DNA

Southern Blot Analysis

Digest Genomic DNA w/ Various Restriction Enzymes

Agarose Gel Electrophoresis and Southern Transfer

Make Non-Radioactive Metacaspase Probe

Hyribidize Probe to Southern Blot

Washes and Chemiluminescent Detection

Data Analysis


Today s objective

Today’s Objective

To Isolate High Quality Genomic DNA from Emiliania huxleyi


Emiliania huxleyi
Emiliania Huxleyi

http://www.soes.soton.ac.uk/staff/tt/

  • Unicellular eukaryotic marine algae

  • Calcium carbonate coccosphere

  • Blooms of > 100,000 sq kilometers



Techniques theoretical basis
Techniques/Theoretical Basis

Breaking open E. huxleyi cells

  • SDS

  • Proteinase K


Techniques theoretical basis1
Techniques/Theoretical Basis

Removal of other Cellular Macromolecules

  • CTAB-cetyltrimethylammonium bromide

  • NaCl

  • Chloroform/isoamyl Extraction


Techniques theoretical basis2
Techniques/Theoretical Basis

Cesium Chloride Gradient

  • Used to obtain highly pure DNA

  • DNA in gradient subjected to centrifugal force of 105,000 xg

  • DNA forms band in gradient at its boyant density


Techniques theoretical basis3
Techniques/Theoretical Basis

Ultraviolet Spectroscopic Analysis of Nucleic Acids

  • Nucleic acids absorb light at 260 nm

  • Proteins absorb light at 280 nm

  • Purity of Nucleic Acid indicated by A260/A280

  • Pure DNA A260/A280 = 1.6-1.8


Next week
Next Week

  • Quantitate amount of DNA isolated

  • Determine the Purity

  • Assess the Integrity

  • Set Up Restriction Digest


ad