DNA Technology and Genomics

Chapter 20

A. P. Biology

Mr. Knowles

Liberty Senior High School

Restriction Endonucleases• Enzymes that recognize specific DNA

sequences in dsDNA (usually 4-6 bp sequence of nucleotides) and digest the DNA at this site.

• Produce fragments of DNA of various lengths. Restriction Fragments.

• Restriction Enzymes isolated from many species of bacteria.

Restriction Enzymes• Several hundred isolated.• Specifically named:

Ex: EcoRI

E = genus

co = species coli

R = strain RY13

I = first endonuclease isolated

Some enzymes...• Others cleave each strand off-center

at specific nucleotides to produce “overhangs”-sticky end cutters. Ex. EcoRI

• Cut cleanly through the DNA at the same position on both strands- blunt end cutters. Ex. Hae III

How Do You Separate DNA Fragments?

• Electrophoresis- the separation of DNA fragments based upon the negative charge of the DNA and its size.

• DNA is negatively charged because of phosphates.

DNA Fingerprinting or Profiling- Many Applications

Genetic Variation• Any two humans have nearly identical

sequences of DNA (99.9%).• But 0.1% is = to 3.0 X 106 nucleotides

(recall the genome is 3.0 X 109 nucletotides).

• DNA profiling takes advantage of polymorphisms (many forms) of repeated sequences in noncoding regions of DNA (near centromeres or ends of chromosomes)

Two Kinds of Polymorphisms• Variable Number Tandem Repeats (VNTR’s) –

9-80 nucleotides that repeat next to each other; the number of times they repeat varies.

• Short Tandem Repeats (STR’s) – 2-8 nucleotides that repeat; also variable in the number of repeats.

• VNTR and STR patterns are inherited from each parent; one set from each parent. Can be used in paternity testing, crimes scene identification, conservation biology, etc.

What’s the point?• Use different restriction enzymes to

make a restriction map- relative positions of sites cut by the enzymes.

• Allows you to recombine DNAs of different organisms-recombinant DNA technology.

• Allows you to identify and match DNAs; DNA Fingerprinting.

Restriction Enzymes Identify Genetic Disease

• Mutations in a gene change the length of the DNA fragments cut by restriction enzymes.

• These different lengths are called Restriction Fragment Length Polymorphisms (RFLPs).

• Fragments can be separated by electrophoresis.

An Application• Glow-in-the-Dark Pets?• http://www.msnbc.msn.com/id/30463427/

http://www.plantsciences.ucdavis.edu/bis101/ANIMATIO/restmap.dir