CHANGING THE LIVING WORLD
How we change the living world…Selective breeding: crossing organisms with desired traits to produce the next generation.
How we change the living world…Hybridization: crossing dissimilar organisms to get the best of both.
How we change the living world…
Inbreeding: continually breeding individuals with similar characteristics.
GENETIC ENGINEERING
Genetic engineering vocab– Recombinant DNA- nucleotide sequences from two
different sources to form a single DNA molecule.
– Transgenic organism – contains a gene from another organism, typically a different species
– Genetically modified organisms (GMOs)- organisms that have acquired one or more genes by artificial means.
Figure 12.1
Genetic Engineering
Genetic engineering: The process of manipulating genes for practical purposes.
Genetic engineering may involve building recombinant DNA
DNA made from two or more different organisms.
Steps in a Genetic Engineering Experiment
Step 1 Isolate Target DNA and plasmid and cut with restriction enzymes
Step 2 Recombinant DNA is produced.
Step 3 Gene cloning: the process by which many copies of the gene of interest are made each time the host cell reproduces.
Step 4 Cells undergo selection and then are screened.
Steps in a Genetic Engineering Experiment
Step 1 The DNA from the organism containing the gene of interest and the vector are cut by restriction enzymes.
A vector is an agent that is used to carry the gene of interest into another cell
Commonly used vectors: viruses, yeast, and plasmids.
circular bacterial DNA
Plasmids
Bacterialchromosome
Remnant ofbacterium
Co
lori
zed
TE
M
Figure 12.7
Plasmid
Bacterial cell
Isolateplasmids.
Figure 12.8-1
Plasmid
Bacterial cell
Isolateplasmids.
DNA
IsolateDNA.
Cell containingthe gene of interest
Figure 12.8-2
Plasmid
Bacterial cell
Isolateplasmids.
DNA
IsolateDNA.
DNA fragmentsfrom cell
Cut both DNAswith sameenzyme.
Gene ofinterest
Othergenes
Cell containingthe gene of interest
Figure 12.8-3
Plasmid
Bacterial cell
Isolateplasmids.
Gene of interest
Recombinant DNA plasmids
DNA
IsolateDNA.
DNA fragmentsfrom cell
Cut both DNAswith sameenzyme.
Gene ofinterest
Othergenes
Mix the DNAs andjoin them together.
Cell containingthe gene of interest
Figure 12.8-4
Plasmid
Bacterial cell
Isolateplasmids.
Recombinant bacteria
Gene of interest
Recombinant DNA plasmids
Bacteria take up recombinant plasmids.
DNA
IsolateDNA.
DNA fragmentsfrom cell
Cut both DNAswith sameenzyme.
Gene ofinterest
Othergenes
Mix the DNAs andjoin them together.
Cell containingthe gene of interest
Figure 12.8-5
Plasmid
Bacterial cell
Isolateplasmids.
Clone the bacteria.
Recombinant bacteriaBacterial clone
Gene of interest
Recombinant DNA plasmids
Bacteria take up recombinant plasmids.
DNA
IsolateDNA.
DNA fragmentsfrom cell
Cut both DNAswith sameenzyme.
Gene ofinterest
Othergenes
Mix the DNAs andjoin them together.
Cell containingthe gene of interest
Figure 12.8-6
Plasmid
Bacterial cell
Isolateplasmids.
Find the clone withthe gene of interest.
Clone the bacteria.
Recombinant bacteriaBacterial clone
Gene of interest
Recombinant DNA plasmids
Bacteria take up recombinant plasmids.
DNA
IsolateDNA.
DNA fragmentsfrom cell
Cut both DNAswith sameenzyme.
Gene ofinterest
Othergenes
Mix the DNAs andjoin them together.
Cell containingthe gene of interest
Figure 12.8-7
Plasmid
Bacterial cell
Isolateplasmids.
Some usesof genes
Gene for pestresistance
Gene fortoxic-cleanupbacteria
Genes may beinserted intoother organisms.
Find the clone withthe gene of interest.
The gene and proteinof interest are isolatedfrom the bacteria.
Clone the bacteria.
Recombinant bacteriaBacterial clone
Gene of interest
Recombinant DNA plasmids
Bacteria take up recombinant plasmids.
Harvestedproteins may beused directly.
Some usesof proteins
Protein for“stone-washing”jeans
DNA
Cell containingthe gene of interest
Protein fordissolvingclots
IsolateDNA.
DNA fragmentsfrom cell
Cut both DNAswith sameenzyme.
Gene ofinterest
Othergenes
Mix the DNAs andjoin them together.
Figure 12.8-8
RESTRICTION ENZYMESmolecular scissors
A Closer Look: Cutting and Pasting DNA with Restriction Enzymes
– Recombinant DNA is produced by combining two ingredients:
A bacterial plasmid The gene of interest
How do we cut them?
• Using restriction enzymes: bacterial enzymes which cut DNA at specific nucleotide sequences
• produce pieces of DNA called restriction fragments.
• Why do you think bacteria contain restriction enzymes?
Restriction Enzymes are palindromes: the same forward as backwards, like RACECAR.
Examples:
GAATTC CCCGGG AAGCTTCTTAAG GGGCCC TTCGAA
G AATTC CCC GGG A AGCTTCTTAA G GGG CCC TTCGA A
Sticky EndsBlunt End
Recognition sequencefor a restriction enzyme
Restrictionenzyme
Sticky
end
Stickyend
DNA
A restriction enzyme cutsthe DNA into fragments.
Figure 12.9-1
Recognition sequencefor a restriction enzyme
Restrictionenzyme
Sticky
end
Stickyend
DNA
A DNA fragment is addedfrom another source.
A restriction enzyme cutsthe DNA into fragments.
Figure 12.9-2
Recognition sequencefor a restriction enzyme
Restrictionenzyme
Sticky
end
Stickyend
DNA
A DNA fragment is addedfrom another source.
A restriction enzyme cutsthe DNA into fragments.
Fragments stick together bybase pairing.
Figure 12.9-3
DNA LIGASE –DNA ligase connects the DNA fragments into one continuous strand (DNA Glue or tape)
Recognition sequencefor a restriction enzyme
Restrictionenzyme
Sticky
end
Stickyend
DNA
DNAligase
Recombinant DNA molecule
A DNA fragment is addedfrom another source.
A restriction enzyme cutsthe DNA into fragments.
Fragments stick together bybase pairing.
DNA ligase joins thefragments into strands.
Figure 12.9-4
Recognition sequences
DNA sequence
Restriction enzyme EcoRI cuts the DNA into fragments.
Sticky end
Your turn to try!!
– Plasmids:• Can easily incorporate foreign DNA
• Are readily taken up by bacterial cells
• Can act as vectors, DNA carriers that move genes from one cell to another
• Are ideal for gene cloning, the production of multiple identical copies of a gene-carrying piece of DNA
Bacterial cells don’t edit the RNA, so how can they make the correct protein?
Genetic Engineers can eliminate the introns from mRNA and reverse the process, producing a DNA strand that is only the instructions for the protein.
Use Reverse Transcriptase
Cell nucleus
DNA ofeukaryoticgene
Test tube
Transcription
Exon Intron Exon ExonIntron
Figure 12.11-1
Cell nucleus
DNA ofeukaryoticgene
RNAtranscript
mRNA
Test tube
Transcription
Introns removed andexons spliced together
Exon Intron Exon ExonIntron
Figure 12.11-2
Cell nucleus
DNA ofeukaryoticgene
RNAtranscript
mRNA
Test tube
Reversetranscriptase
Transcription
Introns removed andexons spliced together
Isolation of mRNA fromcell and addition ofreverse transcriptase
Exon Intron Exon ExonIntron
Figure 12.11-3
Cell nucleus
DNA ofeukaryoticgene
RNAtranscript
mRNA
Test tube
Reversetranscriptase
cDNA strandbeing synthesized
Transcription
Introns removed andexons spliced together
Isolation of mRNA fromcell and addition ofreverse transcriptase
Synthesis of cDNAstrand
Exon Intron Exon ExonIntron
Figure 12.11-4
Cell nucleus
DNA ofeukaryoticgene
RNAtranscript
mRNA
Test tube
cDNA of genewithout introns
Reversetranscriptase
cDNA strandbeing synthesized
Transcription
Introns removed andexons spliced together
Isolation of mRNA fromcell and addition ofreverse transcriptase
Synthesis of cDNAstrand
Synthesis of second DNAstrand by DNA polymerase
Exon Intron Exon ExonIntron
Figure 12.11-5