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DNA Technology
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Copying DNA: PCRCopying DNA: PCR• Polymerase Chain ReactionPolymerase Chain Reaction• Gene Amplification • A method of making many copies of a piece of DNA
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PCR: Copying DNAPCR: Copying DNA
•DNA, nucleotides, buffers, “taq” polymerase, and two primers are placed in a small test tube
• taq polymerasetaq polymerase can work at very high temps
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PCR: Step 1PCR: Step 1
•The DNA is heatedDNA is heated (80oC), the two strands separate
• PrimersPrimers match with complementary bases
•Taq pol. begins adding new nucleotides (5’->3’)
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PCR: Step 2PCR: Step 2
•The tube is cooled the DNA strands together
•Cycle repeated to make several copies
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PCRPCR
Large amounts of DNA can be made Large amounts of DNA can be made from a small starting samplefrom a small starting sample
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Cloning
Just a gene vs. whole organims
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QuickTime™ and aCinepak decompressor
are needed to see this picture.
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Bacterial plasmids in gene cloning
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Cloning a gene• Requires:
– Plasmid (cloning vector), restriction enzymes, and gene of interest
• DNA ligase attaches the sticky ends of DNA fragments together
• Chimeric DNA made!
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Cloning Cloning an an
organismorganism• A body cellbody cell from one organism and an egg cellegg cell from another are fused
• The resulting cell divides divides like a normal like a normal embryoembryo
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Cloning “Dolly”Cloning “Dolly”
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Cutting DNACutting DNA• Restriction enzymesRestriction enzymes cut DNA at specific sequences
• Useful to divide DNA into manageable fragmentsmanageable fragments
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Electrophoresis•DNA can be separated based on size and chargesize and charge
•The phosphate phosphate groupsgroups are negativelynegatively charged
•DNA is placed in a gelgel and electricityelectricity is run through
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Electrophoresis• Negative DNANegative DNA moves toward the positive end
• SmallerSmaller fragments move farther and fasterfarther and faster
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Electrophoresis