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What is recombination
- naturally or artificially change an organism's genome;
- insertion of a gene into a cell
what does recombination result in
- eliminate undesirable phenotypic traits
- combine beneficial traits
- create organisms that can synthesize products needed by humans
What is Biotechnology?
deliberately manipulating an organisms genes and genome
What are the five techniques of genetic engineering
- Multiplying DNA in vitro: (PCR)
- Selecting a clone of recombinant cells
- Separating DNA molecules
- DNA microarrays
- Inserting DNA into cells
Multiplying DNA in vitro: PCR
Polymerase Chain reaction uses polymerase enzymes and changes in temperature to allow for rapid multiplying or replicating of DNA sequences in vitro. provides large quantity of the gene to be used for procedures
Selecting a clone of recombinant cells
marker probes pair with the gene being sought out. markers are radioactive or flourescent
Seperating DNA molecules
- Gel Elecrophoresis is used.
- Southern blot
contain many molecules of DNA or RNA attached to a single carrier such as a glass slide, silicon chips or nylon membranes. This allows scientists to keep lots of genetic material in a single location
Inserting DNA into cells
- Electroporation- elecical current punches holes in cell memrane so DNA can enter
- Gene gun- shoots beads coated with DNA
- Glass pipette- squirted in
When the DNA of both cells combines to form a new molecule recombination has occured
Tools of genetic engineering
- revers transcriptase
- synthetic nucleic acids
- restriction enzymes
- gene libraries
collection of bacterial or virus clones each containing genes of interest. Allows them to go straight to the gene. "gene book"
sequence of nucleotides that can be used to deliver a gene into a cell. Can sometimes be viral genomes, transposons, and plasmids.
- produced by bacterial cells to protect against viruses
- Can cut DNA at specific nucleotide sequences called restriction sites
Synthetic nucleic acids
The enzymes used for DNA replication and RNA transcription can function in the cell (in vivo) and outside the cell (in vitro). Making DNA and RNA in solutions results in the production of synthetic nucleic acids
The use of reverse transcriptase to synthesize cDNA
Reverse Transcriptase is an enzyme that can be used to bond DNA nucleotides to an RNA template (reverse of transcription). This enzyme is produced by retroviruses such as HIV. There are usually multiple copies of mRNA in a cell. Being able to extract them and use them as templates to make DNA is easier than trying to extract and use DNA to make DNA. Reverse transcriptase allows the mRNA to be used to synthesize DNA. DNA produced in this way is called cDNA or complimentary DNA. This DNA lacks introns which makes it easier to insert into prokaryotic cells.
Mutagens are physical and chemical agents that cause mutations or changes in a gene. Mutagens can be deliberately used to mutate genes to gain new variations of a particular gene.
Polymerase chain reaction steps
- 1. "denaturation" the DNA to be replicated is exposed to heat of about 94oC. This breaks the hydrogen bonds and separates the two DNA strands.
- 2. "primed".a mixture of DNA primers, DNA polymerase and lots of the 4 DNA nucleotides (A, G, T and C) is added to the solution containing the separated DNA. When the mixture is cooled to 65oC the two strands of DNA are replicated using the molecules in the mixture that was added.
- 3. called extension and involves raising the temperature back up to 72oC which increases the rate at which DNA polymerase catalyzes the replication of the DNA strands. The end result is lots more pieces of the DNA double helix being replicated.
used to seperate molecues based on their charge, size and shape.
Used to detect DNA for DNA fingerprinting and determine specific strains of infectious organisms
Southern blot is result of transferring DNA from the fragile gel to a sheet of nitrocellulose which is ________ than the electrophoresis gel
Pharmacutical and Theraputic Applications
- Protien synthesis- production of things like insulin
- Genetic screening
- DNA fingerprinting
- Gene therapy
- Medical diagnosis
- Xenotransplants- allow insertion of human genes into other animals to grow organ transplants
- herbicide reistant plants
- salt tolerant plants
- freeze resistant plants
- pest resistant plants
- enhance vitamins