Genetic EngineeringCh 15

“Real World Biology”

Selective Breeding

Selective BreedingPeople select organisms with desired

characteristics to produce next generationTakes advantage of naturally occurring

variation

Selective breeding of teosinte grass by native Americans 6000 years ago led to corn as we now know it

Selective Breeding

HybridizationCross dissimilar organisms to

bring together best of both organismsEx: disease resistance +

increased yield

Benefits include hardier plantsAmerican botanist Luther

Burbank developed more than 800 varieties of plants using selective breeding methods.

Selective Breeding

InbreedingBreeding a line of organisms with similar

characteristicsEx: dog breeds

Risks- decreased genetic variation and increased susceptibility for certain diseases/disordersEx: hip dysplasia

Increasing Variation

Process used to increase the variation normally present in natureBut why?Biotechnology is the

application of a technological process, invention, or method to living organisms.

Increasing variation

Can be accomplished through mutationsMutations are usually random, but can be

induced via radiation and chemical exposurePotential to yield few beneficial mutants with

desirable characteristics not found in original population

Increasing Variation

Bacteria- can treat millions at a time increasing chances of producing useful mutantsEx: oil-digesting bacteria

Increasing Variation

Plants-arresting chromosome separation during meiosis to produce polyploidsKnown to be more vigorous than diploid

relatives

13-2 Manipulating DNA

Mutations are randomHaving a way to alter DNA in a very

specific way to achieve a particular result has huge advantages

Scientists can now use the knowledge of DNA structure and its chemical properties to study and change DNA molecules

Tools of Molecular Biologists

Genetic engineering allows biologists to rewrite the DNA code of an organism

Modern techniques employed canExtracting DNA from cellsCutting it into smaller piecesIdentifying sequences of bases in DNA (genes)Making unlimited copies

Finding Genes

Started with Douglas Prasher (1987) Prasher wanted to find a specific gene in a jellyfish

that codes for a molecule called green fluorescent protein, or GFP

• GFP is a natural protein that absorbs energy from light and makes parts of the jellyfish glow

Prasher thought that GFP from the jellyfish could be linked to a protein when it was being made in a cell

• bit like attaching a light bulb to that molecule

Finding Genes (GFP specifically)

Prasher compared part of the amino acid sequence of the GFP protein to a genetic code tablewas able to predict a probable mRNA base sequence that

would code for this sequence of amino acids

Then used a complementary base sequence to “attract” an mRNA that matched his prediction and would bind to that sequence by base pairing. After screening a genetic “library” with thousands of different

mRNA sequences from the jellyfish, he found one that bound perfectly

Finding Genes

To find the actual gene that produced GFP, Prasher took a gel in which restriction fragments from the jellyfish genome had been separated and found that one of the fragments bound tightly to the mRNAThat fragment contained the actual gene for GFP

This method is called Southern blotting, after its inventor, Edwin Southern.

Finding Genes- Southern Blot Analysis

Finding Genes

Today it is often quicker and less expensive for scientists to search for genes in computer databases where the complete genomes of many organisms are available.

Copying DNA (specific genes)

First step is a polymerase chain reaction (PCR) Heat a piece of DNA

• separates its two strands

DNA cools and added primers bind to the single strands DNA polymerase starts copying the region between the

primers• These copies can serve as templates to make still more

copies.

Polymerase Chain Reaction

Once biologists find a gene, a technique known as polymerase chain reaction (PCR) allows them to make many copies of it.

1. A piece of DNA is heated, which separates its two strands.

Polymerase Chain Reaction

2. At each end of the original piece of DNA, a biologist adds a short piece of DNA that complements a portion of the sequence.

These short pieces are known as primers because they prepare, or prime, a place for DNA polymerase to start working.

Polymerase Chain Reaction

3. DNA polymerase copies the region between the primers. These copies then serve as templates to make more copies.

4. In this way, just a few dozen cycles of replication can produce billions of copies of the DNA between the primers.

Copying DNA

It is relatively easy to extract DNA from cells and tissues.

The extracted DNA can be cut into fragments of manageable size using restriction enzymes.

These restriction fragments can then be separated according to size, using gel electrophoresis or another similar technique

Gel Electrophoresis

Recombinant DNA Technology

It is a form of genetic engineering that cleaves DNA into small fragments and inserts those fragments into a host organismHost may be the same or a different species

Transgenic Organisms

Organisms who have incorporated foreign DNA in their chromosomes and use this new DNA as their own

How to Produce a Transgenic Organism

Step 1: Isolate the gene in the foreign DNA that you want to insertEx: isolate the gene for beta carotene in a

daffodil so you can then add it to rice

Step 2: Cut it out of the chromosome (in daffodil) using restriction enzymes.

Restrictions enzymes are bacterial proteins that have the ability to cut both strands of the DNA molecule at a specific nucleotide sequence

Resulting fragments can have blunt ends or sticky ends

Some Commonly used REs

EcoRI (eco r one)HindIII (hindi three)BamHI (bam h one)TaqI (tack one)

Step 3: Cut host’s DNA with the same RE so cut ends will match up

When DNA from two different organisms joins up- recombinant DNA is formed

VectorsGetting DNA from one organism into

another requires a vectorThe vector introduces the new DNA into the

host cell

Bacterial DNA is often used as a vector

Bacterial DNA

Bacteria contains plasmids- small rings of DNA separate from the bacterium’s larger circular chromosome

The foreign DNA is inserted into the plasmid by cleaving both using the same restriction enzyme

Sticky ends match up and foreign DNA becomes part of plasmid

Gene Cloning

Plasmid with foreign DNA (Now considered recombined DNA) is inserted into a bacterial cell

Plasmids can replicate within the cell and can produce up to 500 copies in the cell

Soon Tons of Copies!

Bacteria clones the recombinant DNAClones-genetically identical copies

How?Bacterial cells themselves will reproduce

quickly, each with hundreds of copies of the recombinant DNA inside (plasmid + foreign DNA)

Introduction into Host Cell

Plasmid is then inserted into a host’s chromosome where it will be replicated each time the cell replicates along with the organism’s other chromosomes

The host cell can transcribe/translate that recombinant DNA into protein just like all other proteins coded in its DNA