Ch. 13.1: BIOTECHNOLOGYObjectives: 1. Explain how the

use of bacteria has contributed to the development of DNA technology.

2. List some recent research trends in recombinant DNA technology.

Vocab: Biotechnology Recombinant DNA

technology

Biotechnology= Use of organisms

to perform practical tasks.

Examples: Use bacteria to make proteins/drug therapies Engineer plants with natural resistance to

insects, drought, fungi, etc. Develop vaccines.

Recombinant DNA

= DNA from 2 different sources combined.

13.2: Bioengineering

Vocab:Plasmid Restriction enzymeGenomic library Nucleic acid probe

Objectives:1. Explain the role of plasmids in

engineering bacteria.2. Explain how biologist “cut and

paste” DNA.3. Describe the procedure used in

cloning a specific gene.4. Identify the usefulness of

recombinant microorganisms.

Bacteria: Work horses of Biotech.

Used to mass produce useful genes + proteins.

Simple organisms 1 chromosome. Plasmids

Reproduce rapidly Easy to manage in a

lab.

Bacteria Plasmids Small, circular DNA Separate from

chromosome. Contains a few genes. Make copies of itself Can be shared/ transferred

b/w bacteria.

Exchange of plasmids is how bacteria build genetic variation.

Genes for antibiotic resistance are shared this way :(

Making Recombinant DNA1. Remove desired gene

from donor cell.2. Remove bacteria

plasmid.3. Insert gene into

plasmid.4. Return plasmid to

bacteria. 5. Gene is transcribed

and translated into protein product.

Recombinant DNAhttp://highered.mcgraw-hill.com/olcweb/cgi/pluginpop.cgi?it=swf::535::535::/sites/dl/free/0072437316/120078/bio38.swf::Early

Restriction Enzymes= Enzymes that CUT foreign DNA sequences.

Each enzyme is cuts a specific sequence (CCCGGG or GAGCT)

Cuts sugar-phosphate backbone of DNA

Make staggered cuts.

Leave “sticky ends” on cuts.

Evolved in bacteria to protect bacteria fr. invading viruses.

Restriction Enzymes: Cut DNA into fragments

Sticky ends = made by staggered cuts; unpaired bases; useful b/c they h-bond w/ complimentary bases in other fragments.

Helps to “sew” fragments together & make recomb. DNA.

BLUNT ends =no staggered cuts; not as useful

Ligase: Glues DNA fragments together

Genetic Cloning = Copies of recomb. DNA (and resulting

proteins) are made by reproducing organisms

Genomic Library= Complete collection of cloned DNA

fragments from an organisms.

When you use restriction enzymes they cut up the donor DNA into MANY fragments.

Each fragment is incorporataed into a plasmid.

You need to figure out which bacteria has desired recomb. Plasmid!

Identifying Desired Recomb. DNA1. Use nucleic acid probe

Radioactively labeled complimentary sequence

(TAGGCT will find and bind to ATCCGA when strands are separated).

2. Insert desired DNA into plasmid sequence for antibiotic resistance.

Recomb. plasmids will lose resistance to antibiotics and will NOT survive when exposed to antibiotic.

Task: Diagram Steps in Creating recombinant DNA clones

Use and illustrate the following key terms…Host DNABacteria PlasmidVectorRestriction EnzymeSticky EndsLigaseBacteria reproduces/plasmid replicationRecombinant DNAClones

Lab: Recombinant Paper PlasmidsGoal: Insert human gene for insulin production into bacteria plasmid.

1. Construct bacteria plasmid. Color code sequences for antibiotic resistance.

2. Label 3’ and 5’ ends of restriction enzymes and plasmid DNA.

3. Identify restriction enzymes that will cut plasmid in sequence for antibiotic resistance. Mark these locations and label the enzyme used.

Lab: Recombinant Paper PlasmidsGoal: Insert human gene for insulin production into bacteria plasmid.

• Label 3’ and 5’ of human DNA

• Determine the enzyme that will allow for removal of human insulin gene AND match up with the sticky ends on the plasmid.

• Create the recombinant plasmid.