GENETIC ENGINEERING

MANIPULATING GENES… Can we make our food taste better? Can we make humans live longer? Can we make X-men like mutants?!? Let’s start with HOW…

3 BILLION BASE PAIRS… How can we make this more manageable… If I wanted to read War & Peace it would be something quite intimidating… But I could make it less scary by breaking it up into small segments…. I will read 20 pages a night! I will be done in a little over two months! Piece of cake…

CUTTING UP DNA: RESTRICTION ENZYMES What are they? Enzymes isolated from certain bacteria strains. They cut DNA at very specific sequences of nucleotides. Hundreds are identified and each cuts at a different sequence

STICKY ENDS Ex: EcoRI restriction enzyme Sequence it recognizes: GAATTC, cuts between G & A Overhanging, single stranded DNA called “sticky” because they could stick back together or to another sticky end

ISOLATING A SPECIFIC GENE Geneticists look for restriction enzymes to cut DNA fragments containing specific genes of interest. GENE Once we have the gene…where do we put it!?

CLONING VECTORS Used to carry a gene from one organism to another. Plasmids: commonly used vectors A small circular piece of DNA, again, coming from bacteria. Separate from prokaryotic DNA, often aid in antibiotic resistance. Origin of replication: where a replication forms to replicate the plasmid

If scientists were to use EcoRI ( or any restriction enzyme) to cut both a segment of DNA and a plasmid vector The sticky ends would be complimentary and could stick together.

TRANSPLANTING GENES Goal: use bacteria to transcribe and translate a DNA sequence to make proteins that some humans cannot make, such as INSULIN 1.Find gene! Cut using restriction enzymes to isolate 2.Make recombinant DNA! Use a plasmid vector to insert gene 3.Cloning DNA- place plasmid vector into a bacterial cell (TRANSFORM) Bacterial cell will then copy plasmid into new cells 4.Expressing the cloned gene: bacteria must successfully transcribe & translate sequence

Recombinant DNA : the result of combining two different DNA sequences that would not normally go together Transgenic Organisms: Inserting a foreign gene into an organism which can then express the gene and pass it on to offspring

DNA FINGERPRINTING Though we are all 99.9% identical we have been able to find the places where we differ in DNA sequence and use these like a fingerprint. In non-coding regions, we all have series of repeats. Ex: GGTTACTAC-GGTTACTAC-GGTTACTAC Will be different on homologous chromosomes… your mother and father’s repeated sequences were most likely different and you have a copy form each.

READING THE FINGERPRINT Read the patterns in each column Where do they match? Matching patterns come form the same person

HOW DO WE GET THAT PICTURE? The method we use for separating fragments of DNA is called GEL ELECTROPHORESIS Use a gel which is porous…

GEL ELECTROPHORESIS What makes the DNA want to move through the gel? CHARGE DNA has a negative (-) charge For charges, opposites attract

SO WHAT DOES IT LOOK LIKE WHEN IT IS DONE? Bands represent different sized fragments of DNA Bigger fragments are closer to the top Smaller fragments travel faster DNA dyed so that it can be seen. Colored dyes and fluorescent dyes are used. Individual segments can be removed and studied

WHAT IF YOU ONLY HAVE A VERY SMALL SAMPLE? We can amplify it, through a method known as… POLYMERASE CHAIN REACTION (PCR) What is polymerase? Where have I heard that word before? Utilizes the efficiency of DNA in replicating itself.

HOW PCR WORKS…

BENEFITS OF PCR Ability to use very small samples crime scenes ancient DNA Can do many trials and experiments on the same DNA from just one sample