CHPT. 20 DNA TECHNOLOGY & GENOMICS & AP LAB #6A

Restriction Enzymes - -found in bacteria, they prevent invasion by foreign bacteria -cut specific sequences -turns out these are VITAL when it comes to manipulating DNA

Restriction Enzymes - restriction site -area where DNA is cut usually only bp’s long -it is a palindrome -some DNA molecules have many of these specific sites… some have none

Naming the R.E.’s - ex. BamH I B = genus Bacillus am = species amyloliquefaciens H = strain (kind) I = order order inwhich this R.E. from this species of bacterium was isolated

Separates DNA DNA segments segments based on size and charge of the molecule

Fragments of DNA move through a “forest of fibers” Small fragments can find the holes and move through the forest more easily… faster

Fragments of DNA move through a “forest of fibers” Large fragments get wrapped around the “trees”, they cannot move as quickly or as far. Large fragments get wrapped around the “trees”, they cannot move as quickly or as far.

Really, it is made like this

Really, it is made like this.

The technique…

The result…

The technique…

The result… #5 is your “marker strand”

Marker Strands… have known bp lengths

How do we “get” the fragments?

In this part of the laboratory, you will use gel electrophoresis to separate samples of DNA that have been digested by. You will then, of unknown size to fragments of a known size to calculate the unknown fragment sizes. In this part of the laboratory, you will use gel electrophoresis to separate samples of DNA that have been digested by restriction enzymes. You will then, compare fragments of unknown size to fragments of a known size to calculate the unknown fragment sizes.

WHY use this type of graph paper?????

How do we “get” the fragments? Better powerpoint on chpt. 20 for this!!!

Result Patterns of DNA markers. DNA fingerprinting Markers are inherited in a Mendelian pattern (Pedigree studies).

Southern Blotting Developed by E.M. Southern in Method to probe DNA for specific RFLP results.

Method Make RFLP fragments. Move fragments from the gel to a nylon sheet. Use probes to make the fragments of interest show up on the sheet.

Method Make RFLP fragments. Move fragments from the gel to a nylon sheet. Use probes to make the fragments of interest show up on the sheet.

Results More permanent record of RFLP results. Very sensitive to small DNA differences.

DNA Technology: Applications 1. Basic Research 2. Medical 3. Forensics 4. Agricultural

DNA Microarray

Future Of DNA Technology Cloning of higher animals. Growth of replacement tissues and organs. Gene therapy to correct DNA defects. ?

Gene Therapy

1) Place gel in chamber 2) Add buffer slowly to each side, let slightly cover the top of the gel (do not have dimples) 3) Use pipet plastic tip to remove any debris 4) Move to the back desk near your power source 5) Plug in DNA near BLACK side 6)Look for bubbles 6)Look for bubbles 7) Watch in awe!!! :-) 7) Watch in awe!!! :-)