chapter 20 dna technology and genomics
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Chapter 20 DNA Technology and Genomics. Chapter 20 DNA Technology and Genomics. DNA cloning DNA probe hybridization Polymerase Chain Reaction (PCR) Gel Electrophoresis Southern Blot DNA sequencing. Chapter 20 DNA Cloning. - PowerPoint PPT PresentationTRANSCRIPT
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Chapter 20DNA Technology and Genomics
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•DNA cloning
•DNA probe hybridization
•Polymerase Chain Reaction (PCR)
•Gel Electrophoresis
•Southern Blot
•DNA sequencing.
Chapter 20DNA Technology and Genomics
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Bacteria have restriction enzymes to attack and destroy invading viral DNA.
Restriction enzymes cut DNA at specific nucleotide sequences leaving “sticky ends.”
DNA ligase can seal these ends, making recombinant DNA.
Chapter 20DNA Cloning
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Restriction fragments can be put into plasmids.
Gene cloning occurs when cells containing these plasmids reproduce.
Genes of interest are marked with a radioactive DNA probe.
Chapter 20DNA Cloning
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If a gene is inserted next to a promoter, the bacteria becomes an expression vector.
Eukaryotic chromosomes allow for bigger segments of DNA.
Eukaryotic cells can also process polypeptides into proteins.
Chapter 20DNA Cloning
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Chopping up the whole genome of an organism produces many DNA fragments containing many genes.
Often, the researcher will save all of them, either in bacteria or in viruses.
These collections of bacterial clones are called genomic libraries.
Chapter 20DNA Cloning
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Chapter 20DNA Probe Hybridization
Samples of DNA
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Chapter 20DNA Probe Hybridization
Heat to 95o to denature
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Chapter 20DNA Probe Hybridization
Introduce radioactive probe
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Chapter 20DNA Probe Hybridization
Allow to cool
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Chapter 20DNA Probe Hybridization
Measure radioactivity
√ x x
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Polymerase chain reaction (PCR) uses DNA polymerase to clone DNA in vitro.
In vitro = in a test tube
In vivo = in a living organism
Chapter 20Polymerase Chain Reaction (PCR)
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DNA FingerprintingHuman DNA contains lots of noncoding sequences that serve no purpose.
This “junk DNA” often repeats over and over.
No two people (except identical twins) have exactly the same repeats.
Chapter 20Gel Electrophoresis
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DNA Fingerprinting
Bill’s chromosome:gene A xxxxx gene B yyy gene C
Bob’s chromosome:gene A xx gene B yyyyy gene C
Chapter 20Gel Electrophoresis
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DNA FingerprintingRestriction enzymes cut DNA at specific places.
Bill’s chromosome:gene A xxxxx gene B yyy gene C
Bob’s chromosome:gene A xx gene B yyyyy gene C
Chapter 20Gel Electrophoresis
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DNA FingerprintingRestriction enzymes cut DNA at specific places.
Bill’s chromosome:gene A xxxxx gene B yyy gene C
Bob’s chromosome:gene A xx gene B yyyyy gene C
Chapter 20Gel Electrophoresis
![Page 17: Chapter 20 DNA Technology and Genomics](https://reader036.vdocument.in/reader036/viewer/2022062500/5681515b550346895dbf7f63/html5/thumbnails/17.jpg)
DNA FingerprintingRestriction enzymes cut DNA at specific places.
Bill’s chromosome:gene A xxxxx gene B yyy gene C
Bob’s chromosome:gene A xx gene B yyyyy gene C
Chapter 20Gel Electrophoresis
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DNA FingerprintingRestriction enzymes cut DNA at specific places.
Bill’s chromosome:
Bob’s chromosome:
Chapter 20Gel Electrophoresis
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DNA FingerprintingLonger fragments travel more slowly through the gel.
Bill’s chromosome:
Bob’s chromosome:
Chapter 20Gel Electrophoresis
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DNA FingerprintingBill: Bob:
Chapter 20Gel Electrophoresis
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DNA FingerprintingBill: Bob:
Chapter 20Gel Electrophoresis
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DNA FingerprintingBill: Bob:
Chapter 20Gel Electrophoresis
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DNA FingerprintingBill: Bob:
Chapter 20Gel Electrophoresis
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Chapter 20Gel Electrophoresis
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The southern blot
1.Do DNA fingerprinting on an entire genome.
2.Blot the DNA from the gel to paper with an alkaline solution. This denatures the DNA.
3.Hybridize with a radioactive probe.
Chapter 20Southern Blot
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The southern blot
Chapter 20Southern Blot
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The southern blot
Chapter 20Southern Blot
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Chapter 20DNA Sequencing
1. Synthesize DNA 3’ → 5’
2. Stop synthesis at random
3. Dideoxyribonuceotides with dye
4. Separate fragments by length
5. Read the colors of the lengths.
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Chapter 20DNA Sequencing
Synthesize DNA 3’ → 5’
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Chapter 20DNA Sequencing
Synthesize DNA 3’ → 5’
5-AGTACCTG-3
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Chapter 20DNA Sequencing
Synthesize DNA 3’ → 5’
5-AGTACCTG-3 3-TCATGGAC-5
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Chapter 20DNA Sequencing
Stop the process with didexoyribonucleotides
5-AGTACCTG-3 3-TCATGGAC-5
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Chapter 20DNA Sequencing
Stop the process with didexoyribonucleotides
5-AGTACCTG-3 3-TCATG
![Page 34: Chapter 20 DNA Technology and Genomics](https://reader036.vdocument.in/reader036/viewer/2022062500/5681515b550346895dbf7f63/html5/thumbnails/34.jpg)
Chapter 20DNA Sequencing
Add a different dye to each didexoyribonucleotide
A T C G
5-AGTACCTG-3
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Chapter 20DNA Sequencing
Stop the synthesis at random times with different dyes
A T C G
5-AGTACCTG-3
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Chapter 20DNA Sequencing
Stop the synthesis at random times with different dyes
5-AGTACCTG-3 3-TCATGG
![Page 37: Chapter 20 DNA Technology and Genomics](https://reader036.vdocument.in/reader036/viewer/2022062500/5681515b550346895dbf7f63/html5/thumbnails/37.jpg)
Chapter 20DNA Sequencing
Stop the synthesis at random times with different dyes
5-AGTACCTG-3 3-TCATGG 3-T
![Page 38: Chapter 20 DNA Technology and Genomics](https://reader036.vdocument.in/reader036/viewer/2022062500/5681515b550346895dbf7f63/html5/thumbnails/38.jpg)
Chapter 20DNA Sequencing
Stop the synthesis at random times with different dyes
5-AGTACCTG-3 3-TCATGG 3-T 3-TCATGGAC
![Page 39: Chapter 20 DNA Technology and Genomics](https://reader036.vdocument.in/reader036/viewer/2022062500/5681515b550346895dbf7f63/html5/thumbnails/39.jpg)
Chapter 20DNA Sequencing
Stop the synthesis at random times with different dyes
5-AGTACCTG-3 3-TCATGG 3-T 3-TCATGGAC 3-TCA
![Page 40: Chapter 20 DNA Technology and Genomics](https://reader036.vdocument.in/reader036/viewer/2022062500/5681515b550346895dbf7f63/html5/thumbnails/40.jpg)
Chapter 20DNA Sequencing
Stop the synthesis at random times with different dyes
5-AGTACCTG-3 3-TCATGG 3-T 3-TCATGGAC 3-TCA 3-TCATG
![Page 41: Chapter 20 DNA Technology and Genomics](https://reader036.vdocument.in/reader036/viewer/2022062500/5681515b550346895dbf7f63/html5/thumbnails/41.jpg)
Chapter 20DNA Sequencing
Stop the synthesis at random times with different dyes
5-AGTACCTG-3 3-TCATGG 3-T 3-TCATGGAC 3-TCA 3-TCATG 3-TCA T
![Page 42: Chapter 20 DNA Technology and Genomics](https://reader036.vdocument.in/reader036/viewer/2022062500/5681515b550346895dbf7f63/html5/thumbnails/42.jpg)
Chapter 20DNA Sequencing
5-AGTACCTG-3 3-TCATGG 3-T 3-TCATGGAC 3-TCA 3-TCATG 3-TCA T
Arrange the fragments by length
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Chapter 20DNA Sequencing
3-T 3-TC 3-TCA 3-TCA T 3-TCATG 3-TCATGG 3-TCATGGA3-TCATGGAC
Arrange the fragments by length
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Chapter 20DNA Sequencing
3-T 3-TC 3-TCA 3-TCA T 3-TCATG 3-TCATGG 3-TCATGGA3-TCATGGAC
Read the colors
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Chapter 20DNA Sequencing
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Chapter 20DNA Sequencing
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RFLPs (“RIF-lips”), or restriction fragment length polymorphisms, are differences in homologous chromosomes that give different length restriction fragments.
Chromosome walking means finding where fragments of DNA overlapped in the genome.
Chapter 20DNA Technology and Genomics
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Genomics is the systematic study of entire genomes.
Proteomics is the study of all the proteins encoded by a genome.
Single nucleotide polymorphisms (SNPs) are useful markers for studying variation.
Chapter 20DNA Technology and Genomics
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Uses of DNA Technology:
Testing for genetic diseases
Large scale production of drugs
Gene therapy
Forensics
Genetic engineering.
.
Chapter 20DNA Technology and Genomics