RECOMBINANT DNAIntroduction

Tools of Genetics: Recombinant DNA and Cloning• The New Genetics pp. 38-39• Summarize:

• How do scientists move genes from one organism to another? • Explain the role of restriction enzymes• Explain how the DNA fragments are combined• Explain the role of cloning in the making of insulin

Tools of Genetics

If scientists want to separate the gene for insulin, what tools would they use?

Tools of Genetics

• After the insulin gene is isolated, what is the next step?

Tools of Genetics

What is E.coli?

Why and how is it used?

Tools of Genetics

• What is this step?

• What is needed to make it happen?

Recombinant DNA (rDNA)

• Method to create a new DNA molecule by piecing together different DNA molecules

• When cells accept the rDNA, they will “express” the new genes by making the new proteins• Cells are “genetically engineered”

A summary of the process:

• Mechanism of Recombination – YouTube

• Recombinant DNA ©2004 Demonstratives, Inc. - YouTube

Genetically engineered products- using recombinant DNA (rDNA)Why?

Products in the body are either:

(1) Made in too small quantities

(2) Made at the wrong time

(3) Lack an important characteristic

(4) Can be made in greater quantity for medical use

Uses rDNA – proteins made by body in small quantities

• Insulin (first rDNA product; 1982)• Growth hormone

• Children with insufficient growth hormone or poor kidney function

• Blood clot prevention (plasminogen activator)• Larger quantities used to prevent heart attacks or strokes

• Blood clotting factor – for hemophilia• Gamma interferon – fight cancer growth

Genetic Engineering –Recombinant DNA How?

• Identify a molecule produced by a living organism

• Isolate the instructions (DNA sequence = gene)

• Put the instructions into another cell or organism

• Allow the cell to replicate • Harvest the desired product

What do we need to learn about to understand how recombinant DNA products?

• Genes and proteins• Bacteria

• Plasmids – extra ring of DNA

• Copying cells - Mitosis• How are enough protein is made? PCR

A more detailed explanation• http://

www.youtube.com/watch?v=GNMJBMtKKWU&feature=related

• Plasmids | Genetics | Biology - YouTube

The Vector (E. coli bacteria).

Chromosome & Plasmid

Restriction Enzymes• Let us analyze the words

• “Restriction endonuclease”

What details is this name telling us?

• “DNA Ligase”:

Restriction Enzyme

Restriction map:

“When scientists study a DNA molecule, one of the first things they do is to figure out where the restriction sites are. They then create a restriction map, showing the locations of cleavage sites for many different enzymes.”

Lambda DNA –

Lambda DNA genome: 48,502 base pairs

21

Lambda DNA Sequence• CTGTCGTTTCCTTTCTCTGTTTTTGTCCGTGGAATGAACAATGGAAGTCA ACAAAAAGCAGCTGGCTGACATTTTCGGTGCGAGTATCCGTACCATTCAG

AACTGGCAGGAACAGGGAATGCCCGTTCTGCGAGGCGGTGGCAAGGGTAA TGAGGTGCTTTATGACTCTGCCGCCGTCATAAAATGGTATGCCGAAAGGG ATGCTGAAATTGAGAACGAAAAGCTGCGCCGGGAGGTTGAAGAACTGCGG CAGGCCAGCGAGGCAGATCTCCAGCCAGGAACTATTGAGTACGAACGCCA TCGACTTACGCGTGCGCAGGCCGACGCACAGGAACTGAAGAATGCCAGAG ACTCCGCTGAAGTGGTGGAAACCGCATTCTGTACTTTCGTGCTGTCGCGG ATCGCAGGTGAAATTGCCAGTATTCTCGACGGGCTCCCCCTGTCGGTGCA GCGGCGTTTTCCGGAACTGGAAAACCGACATGTTGATTTCCTGAAACGGG ATATCATCAAAGCCATGAACAAAGCAGCCGCGCTGGATGAACTGATACCG GGGTTGCTGAGTGAATATATCGAACAGTCAGGTTAACAGGCTGCGGCATT TTGTCCGCGCCGGGCTTCGCTCACTGTTCAGGCCGGAGCCACAGACCGCC GTTGAATGGGCGGATGCTAATTACTATCTCCCGAAAGAATCCGCATACCA GGAAGGGCGCTGGGAAACACTGCCCTTTCAGCGGGCCATCATGAATGCGA TGGGCAGCGACTACATCCGTGAGGTGAATGTGGTGAAGTCTGCCCGTGTC GGTTATTCCAAAATGCTGCTGGGTGTTTATGCCTACTTTATAGAGCATAA GCAGCGCAACACCCTTATCTGGTTGCCGACGGATGGTGATGCCGAGAACT TTATGAAAACCCACGTTGAGCCGACTATTCGTGATATTCCGTCGCTGCTG GCGCTGGCCCCGTGGTATGGCAAAAAGCACCGGGATAACACGCTCACCAT GAAGCGTTTCACTAATGGGCGTGGCTTCTGGTGCCTGGGCGGTAAAGCGG CAAAAAACTACCGTGAAAAGTCGGTGGATGTGGCGGGTTATGATGAACTT GCTGCTTTTGATGATGATATTGAACAGGAAGGCTCTCCGACGTTCCTGGG TGACAAGCGTATTGAAGGCTCGGTCTGGCCAAAGTCCATCCGTGGCTCCA CGCCAAAAGTGAGAGGCACCTGTCAGATTGAGCGTGCAGCCAGTGAATCC CCGCATTTTATGCGTTTTCATGTTGCCTGCCCGCATTGCGGGGAGGAGCA GTATCTTAAATTTGGCGACAAAGAGACGCCGTTTGGCCTCAAATGGACGC CGGATGACCCCTCCAGCGTGTTTTATCTCTGCGAGCATAATGCCTGCGTC ATCCGCCAGCAGGAGCTGGACTTTACTGATGCCCGTTATATCTGCGAAAA GACCGGGATCTGGACCCGTGATGGCATTCTCTGGTTTTCGTCATCCGGTG AAGAGATTGAGCCACCTGACAGTGTGACCTTTCACATCTGGACAGCGTAC AGCCCGTTCACCACCTGGGTGCAGATTGTCAAAGACTGGATGAAAACGAA AGGGGATACGGGAAAACGTAAAACCTTCGTAAACACCACGCTCGGTGAGA CGTGGGAGGCGAAAATTGGCGAACGTCCGGATGCTGAAGTGATGGCAGAG CGGAAAGAGCATTATTCAGCGCCCGTTCCTGACCGTGTGGCTTACCTGAC CGCCGGTATCGACTCCCAGCTGGACCGCTACGAAATGCGCGTATGGGGAT GGGGGCCGGGTGAGGAAAGCTGGCTGATTGACCGGCAGATTATTATGGGC CGCCACGACGATGAACAGACGCTGCTGCGTGTGGATGAGGCCATCAATAA AACCTATACCCGCCGGAATGGTGCAGAAATGTCGATATCCCGTATCTGCT GGGATACTGGCGGGATTGACCCGACCATTGTGTATGAACGCTCGAAAAAA CATGGGCTGTTCCGGGTGATCCCCATTAAAGGGGCATCCGTCTACGGAAA GCCGGTGGCCAGCATGCCACGTAAGCGAAACAAAAACGGGGTTTACCTTA CCGAAATCGGTACGGATACCGCGAAAGAGCAGATTTATAACCGCTTCACA CTGACGCCGGAAGGGGATGAACCGCTTCCCGGTGCCGTTCACTTCCCGAA TAACCCGGATATTTTTGATCTGACCGAAGCGCAGCAGCTGACTGCTGAAG AGCAGGTCGAAAAATGGGTGGATGGCAGGAAAAAAATACTGTGGGACAGC AAAAAGCGACGCAATGAGGCACTCGACTGCTTCGTTTATGCGCTGGCGGC GCTGCGCATCAGTATTTCCCGCTGGCAGCTGGATCTCAGTGCGCTGCTGG CGAGCCTGCAGGAAGAGGATGGTGCAGCAACCAACAAGAAAACACTGGCA GATTACGCCCGTGCCTTATCCGGAGAGGATGAATGACGCGACAGGAAGAA CTTGCCGCTGCCCGTGCGGCACTGCATGACCTGATGACAGGTAAACGGGT GGCAACAGTACAGAAAGACGGACGAAGGGTGGAGTTTACGGCCACTTCCG TGTCTGACCTGAAAAAATATATTGCAGAGCTGGAAGTGCAGACCGGCATG ACACAGCGACGCAGGGGACCTGCAGGATTTTATGTATGAAAACGCCCACC ATTCCCACCCTTCTGGGGCCGGACGGCATGACATCGCTGCGCGAATATGC CGGTTATCACGGCGGTGGCAGCGGATTTGGAGGGCAGTTGCGGTCGTGGA ACCCACCGAGTGAAAGTGTGGATGCAGCCCTGTTGCCCAACTTTACCCGT GGCAATGCCCGCGCAGACGATCTGGTACGCAATAACGGCTATGCCGCCAA CGCCATCCAGCTGCATCAGGATCATATCGTCGGGTCTTTTTTCCGGCTCA GTCATCGCCCAAGCTGGCGCTATCTGGGCATCGGGGAGGAAGAAGCCCGT GCCTTTTCCCGCGAGGTTGAAGCGGCATGGAAAGAGTTTGCCGAGGATGA CTGCTGCTGCATTGACGTTGAGCGAAAACGCACGTTTACCATGATGATTC GGGAAGGTGTGGCCATGCACGCCTTTAACGGTGAACTGTTCGTTCAGGCC ACCTGGGATACCAGTTCGTCGCGGCTTTTCCGGACACAGTTCCGGATGGT CAGCCCGAAGCGCATCAGCAACCCGAACAATACCGGCGACAGCCGGAACT GCCGTGCCGGTGTGCAGATTAATGACAGCGGTGCGGCGCTGGGATATTAC GTCAGCGAGGACGGGTATCCTGGCTGGATGCCGCAGAAATGGACATGGAT

• And 10 pages more

If we are using the restriction enzymes