Difference between biotechnology and genetic engineering • Biotechnology – the use of technology to modify organisms, cells, and their molecules to achieve practical benefits • Genetic engineering – the manipulation of organisms’ genetic material by adding, or deleting, or transplanting genes from one organism to another. Transgenics: genes from another sperices is inserted AGRICULTURE Already changing our world significantly. • Pest- and disease-resistant crops • Dramatically higher crop yields • Foods with enhanced nutrition HUMAN HEALTH Tremendous potential, but limited success so far. • Improved treatment of disease through more effective medicine • Improved diagnosis and screening for genetic diseases FORENSIC SCIENCE Advances already improving the justice system. • Improved capabilities of law enforcement • Important reforms to the criminal justice system
CHOPup DNA from a donor species that exhibits a trait of interest. AMPLIFY small samples of DNA into more useful quantities. INSERT pieces of DNA into bacterial cells or viruses. GROWseparate colonies of bacteria or viruses, each containing some donor DNA. Not all of the tools are used in all biotechnology applications—some use only one or a few of these techniques.
Chop: Isolating a Gene of Interest Using Restriction Enzymes The gene of interest is located on a section of DNA from the donor species. 1 Gene of interest Source DNA Restriction introduce restriction enzymes that target a particular base-pair sequence on either side of the gene. 2 Restriction enzymes In this example, the restriction enzyme recognizes the sequence ATCGAT and cuts between the first A and T. The restriction enzymes bind to their target base-pair sequence and cut the strand of DNA. 3 The gene of interest has now been separated from the donor’s DNA. 4
Amplify: Polymerase Chain Reaction (PCR) A solution containing an isolated segment of DNA is heated, separating the double-stranded DNA into two-single strands. 1 Heat DNA polymerase Primer The enzyme DNA polymerase is added along with primers and a large number of free nucleotides, and the solution containing the segments is cooled. 2 Cool Free nucleotides 3 DNA polymerase adds complementary bases to each single strand. 4 The result is two identical copies of the original segment of DNA. This process can be repeated again and again until there are billions of identical copies of the target sequence.
Insert: Using Plasmids to Transfer DNA from One Organism to Another A target segment of source DNA is isolated using a restriction enzymes. Using the same restriction enzyme, a single cut is made in a bacterial plasmid. 1 Source cell Bacterial cell Plasmid Gene of interest The two segments now share complementary bases at their ends and fit perfectly together. 2 3 The plasmid—now including the gene of interest—is inserted back into the bacterial cellwhere it can be expressed and replicated.
Grow: Creating a Gene Library To create a gene library, a large amount of DNA is chopped up using restriction enzymes. 1 Gene A Gene B Gene C 2 Each piece is inserted into a plasmid, and each type of plasmid is introduced into a different bacterial cell. Plasmid A Plasmid B Plasmid C The bacteria are allowed to divide repeatedly, each producing a clone of the foreign DNA fragment it carries. 3 Bacterium A Bacterium B Bacterium C GENE LIBRARY Together, all of the different bacterial cells contain all of the different fragments of the original DNA. 4
At the cutting edge of biotech, CRISPR is a tool with the potential to revolutionize medicine. • CRISPR • clustered regularly interspaced short palindromic repeats • system for editing DNA with a great deal of precision and efficiency • enables researchers to modify almost any gene in any organism • naturally occurs in almost half of all bacteria as a mechanism for recording encounters with viral DNA and using that information to protect against future infections
How CRISPR Works CRISPR is a gene editing system that brings greater precision and efficiency to modifying genomes. After identifying a particular DNA sequence of interest, researchers synthesize an RNA “guide” molecule with a sequence that matches the target gene to be sliced. 1 DNA sequence of interest RNA guide The sequences for the CRISPR RNA and Cas9 enzyme are introduced to target cells using a plasmid. 2 Plasmid RNA sequence Cas9 sequence Within the cell, the RNA leads the Cas9 enzyme to exactly the desired location on the DNA, and Cas9 cuts the DNA there. 3 Cas9 enzyme New DNA sequence At the location where the DNA is cut, a sequence can be inserted that repairs or alters the host cell’s DNA. 4
Mosquito DNA can be altered to block malaria from spreading. Potential of Crispr Risks of Crispr • Legal issues surrounding who invented it/who can profit from it • Ethical issues • Potential for editing human embryos, editing sperm and egg • Difficult to predict consequences of introducing altered genes into genomes of natural populations Human trials are under way in which CRISPR-altered white blood cells (in blue) are being used to fight cancer cells (red). With CRISPR technology, beagles have been engineered to be born with twice as much muscle as a typical dog.
Biotechnology can improve food nutrition and make farming more efficient and eco-friendly. Ancestral corn • Genetic engineering – the manipulation of a species’ genome in ways that do not normally occur in nature • Recombinant DNA technology has sped up this process, enabling the combination of DNA from two or more sources into a product. Modern corn For many generations, farmers have carefully selected for the most desirable characteristics in corn, resulting in larger, juicier kernels.
Almost everyone in the United States uses genetically modified crops regularly without knowing it. • Two factors driving adoption of modified crops in the U.S.: • Plants with insecticides engineered into them • Plants with herbicide-resistant genes engineered into them 14% 7% 7% 86% 93% 93% Corn Cotton Soybeans Proportion of crops that are not genetically modified Proportion of crops that are genetically modified
More GM foods Flavr – Savr tomato The first GM crop approved by FDA for human consumption Golden Rice – rice with daffodil genes for production of Vit A
FDA approved two GM foods (Mar 20, 2015) Arctic apple and Innate potatoes Conventional apple: Browning Arctic apple: No browning Conventional potato: Browning More acrylamide Innate potato: Fewer black spots Less acrylamide
Insect Resistance Corn engineered to contain spores of the bacterium Bacillus thuringiensis (Bt) kills insect pests but does not harm humans. Bt crystal gene Bacterium (Bacillus thuringiensis) Corn genome Corn cell Corn plant destroyed by butterfly larvae (caterpillars). Bacterial gene coding for Bt crystals, which are poisonous to the caterpillars, is inserted directly into the corn plant’s DNA. Bt crystals within corn cells prevent insect predation, so pesticides are no longer needed. 1 2 3
The larger salmon carries a growth hormone gene that keeps it growing year-round rather than in the summer only. Herbicide Resistance Faster Growth and Bigger Bodies Roundup Ready corn, soybeans, cotton, sugar beets etc
Bt corn is ____ A. Resistant to insecticides B. Resistant to herbicides C. Resistant to insects D. Resistant to low temperatures
Genetically modified bacteria - Insulin - Human growth hormone - Erythropoetin - Blood Clotting protein
what are the possible risks of genetically modified foods? “Organisms that we want to kill may become invincible.” “Organisms that we don’t want to kill may be killed inadvertently.” “Genetically modified crops are not tested or regulated adequately.” “Eating genetically modified foods is dangerous.” “Loss of genetic diversity among crop plants is risky.” “Hidden costs may reduce the financial advantages of genetically modified crops.” Featherless birds are cheaper for farmers and consumers. But there are unintended consequences, including vulnerability to mosquitoes and other parasites.
A goat which produces human blood proteins in its milk is ___ A. Genetically modified but not transgenic B. Genetically modified and transgenic C. Not genetically modified and not transgenic D. Not genetically modified but transgenic
DNA applications • Biotechnology has the potential to • Bring about Social justice • Better human health Forensics Paternity Wildlife biology Systematics Genetic Testing
Current news: Criminals identified through genetic genealogy databases. Golden state killer caught in April 2018
The first reported application of plant DNA evidence involved the molecular identification of seed pods from a Palo Verde tree. It was used to link a suspect to a particular crime scene in an investigation called the Bogan case in 1995
You need DNA as starting materialSample size has decreased over time Touch DNA method: as it analyzes skin cells gathered from the crime scene Size of a quarter..1980s Size of a dime…..1990s If you can see it, you can analyze it Just 7/8 skin cells….2003
DNA is an individual identifier: the uses and abuses of DNA fingerprinting. • Convicted based on DNA evidence left at crime scene Colin Pitchfork was the first criminal brought to justice because of DNA fingerprinting.
Biotechnology in Forensics Short tandem repeats, or STRs, are sequences of DNA (commonly 4 or 5 nucleotides) that repeat over and over again. They occur in some of the most highly variable regions of an individual’s DNA. • Approximately 99.9% of the DNA sequences of two individuals are the same. • There are still approximately three million base-pair differences between two people. • Identical twins are the exception. • STRs (short tandem repeats) are used to determine someone’s genetic fingerprint. INDIVIDUAL A Short tandem repeat (STR) NUMBER OF REPEATS Chromosome from mother 3 Chromosome from father 14 3 / 14 STR region Individual A’s alleles for this STR region INDIVIDUAL B Chromosome from mother 5 Chromosome from father 11 5 / 11 Individual B’s alleles for this STR locus For a given region, the STR sequence is the same, but its number of repeats differs.
Your Unique Identifier: DNA Fingerprint 2 SORT THE FRAGMENTS BY SIZE Amplified DNA fragments are poured into an electrophoresis gel and an electrical charge is applied. Because DNA is a negatively charged molecule, the fragments move toward the positively charged electrode. Smaller pieces (having fewer repeats) move across the gel more quickly than larger pieces. A DNA fingerprint is created by determining which alleles an individual carries for 13 different STR regions. • Useful for comparing a suspect’s DNA to DNA found at a crime scene • Amplify the STR region by PCR • Sort fragments by gel electrophoresis • Not totally foolproof 1 AMPLIFY THE STR REGION For each of the 13 STR regions used, the DNA fragment is amplified using PCR, resulting in huge numbers of those fragments. The fragments differ in size depending on how many times the repeating unit of that STR is repeated. Individual A’s DNA sample Direction of DNA strand movement Electrical charge
Is Steve likely to be the father? Steve M= Mother’s DNA C= Child’s DNA A. Yes B. No
DNA profiling/DNA Fingerprinting/DNA typing (comparisons of DNA samples) Obtain DNA samples Amplify STR regions by PCR Cut with specific restriction enzyme Separate fragments on gel Compare patterns Note: DNA profiling is NOT DNA sequencing…
The pattern of bars in a DNA fingerprint shows….. A. The order of bases in a particular gene B. The presence of various sized fragments from chopped up DNA C. The order of genes along particular chromosomes D. The exact location of a specific gene in a genomic library
Biotechnology and health • Making useful medical products • Gene therapy • Personalized medicine • Personal DNA analysis • Measuring Exposomes (using biotech to measure all the exposures of an individual and how those exposures relate to health)
The treatment of diseases and production of medicines are improved with biotechnology. • Prevent, Cure diseasesand Treat diseases • The treatment of diabetes • Insulin produced using recombinant DNA technology • Human growth hormone (HGH) • Can now be produced by transgenic bacteria • Erythropoietin • Can now be produced from cells derived from hamster ovaries
Why has gene therapy had such a poor record of success in curing diseases? • Difficulty getting the working gene into the specific cells where it is needed • Difficulty getting the working gene into enough cells and at the right rate to have a physiological effect • Difficulty arising from the transferred gene getting into unintended cells • Difficulty regulating gene expression
Personal DNA analysis 23andMe Company is Back on Track after Meeting FDA Requirements: Oct 21, 2015 Using a saliva sample for DNA analysis Genetic Markers -used to discover disease genes http://www.youtube.com/watch?v=F7UHBBHhoj0
Biotechnology and health All the environmental factors that one is exposed to
Personalized medicine uses different OMICS fields to come up with an individualized treatment Integrated Personal Omics Different OMICS field • Genomics (data on DNA : genome) • Proteomics (data on proteins: Proteome) • Metabolomics (data on small molecules (the metabolome) • Transcriptomics (data on mRNA transcripts; the transcriptome) • Epigenomics (experiences changing gene expressions) • Microbiome