DNA Technology Ch. 20

The Human Genome The human genome has over 3 billion base pairs 97% does not code for proteins Called “Junk DNA” or “Noncoding DNA” Can control gene expression, but most are repetitive segments that aren’t transcribed Recombinant DNA is when DNA from two different sources are combined This happens all the time in nature: Viral Transduction Bacterial Transformation Conjugation Transposons

Scientists use recombinant DNA techniques in genetic engineering Some uses of recombinant DNA: Produce proteins (ex. Human growth hormone) Prepare lots of copies of a gene to analyze Create novel organisms Gene Therapy Replacing a nonfunctioning gene with a functioning one Genes then entered into body So far limited success

Restriction Enzymes Restriction enzymes cut DNA at specific recognition sites These cuts are typically staggered, which creates single- stranded sticky ends These pieces created by restriction enzymes are called restriction fragments Some common restriction enzymes are: EcoRI BamHI HindIII

Gene Cloning A desired gene is isolated The gene is then inserted into a plasmid The plasmid is inserted into a vector (cell that can carry plasmid, like bacteria) Cloning the gene by allowing the vector to reproduce by fission. As the vector reproduces asexually it is also creating copies of the gene. Millions of copies of the gene are produced Identify the bacteria that have the gene Extract the genes

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Gel Electrophoresis Separates DNA by size based on their rate of movement through agarose gel in an electric field The smaller the molecule, the faster it moves DNA has a negative charge and is attracted to the positive side of the gel The DNA is first cut with restriction enzymes before being run in the gel Useful in several ways Comparing DNA sample with other samples (criminal cases) You can use a DNA probe to identify the location of certain DNA sequences DNA probe: radioactively labeled single strand of DNA used to find a sequence of DNA

Restriction Fragment Length Polymorphisms Usually referred to as “RFLPs” Noncoding regions of human DNA have restriction fragment pattern that is different in every individual These differences are called RFLPs RFLP analysis of an individuals DNA gives a DNA fingerprint Everybody’s RFLPs are unique, except for identical twins

Southern Blot Technique

Polymerase Chain Reaction Technique that takes a piece of DNA and amplifies it (quickly makes lots of copies) You can make billions of copies of a DNA fragment in hours DNA is incubated in a test tube with: Special type of DNA polymerase Nucleotides DNA Primers

Complementary DNA (cDNA) Scientists clone genes in bacteria Bacteria don’t have a way of cutting out introns so in order to clone a gene in a bacteria you must have a gene with no introns Scientists use mRNA and reverse transcriptase (taken from retroviruses) to make DNA from the RNA. This DNA carries the coding sequence, but without the introns because it came from mRNA