Chapter 5B: Labeled Tracers and Nucleic Acid Hybridization

Why use radioactive tracers?

To detect very small amounts of moleules

What is autoradiography?

Detecting radioactive molecules using X-ray film

Advantage of nonradioactive tracers?

Safer and no radioactive waste

How does enzyme amplification work?

Enzyme on probe makes many product molecules 

What is chemiluminescence?

Product emits light to amplify signal 

What is the goal of Southern blotting? 

Detect specific DNA sequences

Why not stain DNA directly?

Too many fragments = blurred streak

Main steps of southern blotting?

1. Gel

2. Transfer

3. Denature

4. Hybridize

5. Detect 

Why denature DNA?

Only single-stranded DNA hybridizes with probe

What is the purpose of probe?

Binds complementary sequences to reveal DNA fragments

What are mini satellites? 

Short repeated DNA sequences

Why are DNA fingerprints unique?

Repeat numbers differ between people

Uses of DNA fingerprinting?

Forensics, paternity, population genetics

Why use Haelll?

Cutes outside minisatellites, leaving repeats intact

What is DNA typing? 

Using probes for specific loci → simpler patterns 

How many probes needed to ID someone?

4-5 different probes

How is paternity tested? 

Compare childs bands with parents bands 

How is forensic identification done?

match crime scene DNA to suspect

How sensitive is autoradiography? 

Detects very tiny amounts (picograms) 

What is a polycistronic message?

Pme mRNA encoding multiple genes (from lac operon example)

Why use a nitrocellulose or nylon membrane? 

Creates a permanent copy of DNA pattern 

How does a probe bind to DNA? 

Base-pairing with complementary sequence

What does “signal amplification” mean?

Small target = large detectable signal