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Week 10 Notes- Bio

by: Raquel Notetaker

Week 10 Notes- Bio BIOL 1010

Raquel Notetaker
GPA 3.5

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2.8 DNA Technologies & Genomics 2.9 New frontiers: Personalized Medicine. Hope or Hype?
Introduction to Biology
Class Notes
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This 48 page Class Notes was uploaded by Raquel Notetaker on Tuesday April 12, 2016. The Class Notes belongs to BIOL 1010 at Rensselaer Polytechnic Institute taught by in Spring 2016. Since its upload, it has received 17 views. For similar materials see Introduction to Biology in Biology at Rensselaer Polytechnic Institute.


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Date Created: 04/12/16
DNA T echnologies and Society What do these pictures have in common? Video on therapeutic proteins Video Moving your gene of interest into a vector or carrier 12.2 Enzymes are used to “cut and paste” DNA Animation: Restriction Enzymes Copyright © 2009Pearson Education, Inc. DNA restriction endonucleases- tools of the trade for molecular technologies From cloned gene to new host and product E. coli bacterium Plasmid Cell with DNA 1 Isolate containinggene Bacterial plasmid of interest chromosome 2 Isolate DNA 3 Cut plasmid DNA with enzyme Geneof interest 4 Cut cell’s DNA with sameenzyme Gene of interest 5 Combinetargetedfragment and plasmid DNA Examplesof geneuse 6 AddDNA ligase, which closes thecircle with covalentbonds Genesmay beinserted Recombinant into otherorganisms DNA Gene plasmid of interest 7 Putplasmid 9 Genesor proteins are isolated fromthe into bacterium clonedbacterium bytransformation Recombinant bacterium Harvested proteins Examplesof 8 Allow bacterium maybe proteinuse to reproduce useddirectly Animation: Cloning a Gene Clone of cells Eukaryotic genes in prokaryotes cause problems Making cDNA • McGraw-Hill Online Learning Center • Play Animation: cDNA • If you want a protein product and the gene is cloned in a bacterium you need to use a cDNA • Or clone your gene into a eukaryotic cell CRISPR T echnology Many bacteria contain an interesting DNA-based immune system, called CRISPR.(clustered, regularly interspaced short palindromic repeats ). This system consists of a set of DNA-editing enzymes and an array of repeated sequences interspersed with DNA that is derived from pathogens that the bacterium or its ancestors have been infected with. When these pathogens show up in the cell, these sequences are used to target the pathogenic DNA for destruction. Scientists are exploiting this system to make very precise changes in DNA of a variety of organisms. VIDEO Typical CRISPR region in bacterial genome. Repeats are important for the functioning of the system; the spacers contain DNA sequences derived from pathogens. The cas genes are concerned with editing bacterial DNA and cutting foreign DNA. Interference or Defense Phase of CRISPR System Adaptation Phase of CRISP System Attendance exercise • What would you need to engineer the genes for human growth hormone into E. coli? You do not need to know exact details but explain in a brief narrative. Post answers to the bulletin board. CSI-1010 Discovery video: DNA Forensics DNA Profiling Need to amplify the sample DNA • Some samples are very small and the particular region of interest is a small portion of the DNA. A single hair has been sufficient to identify a suspect. • If we copy variable regions of the genome, we would have more DNA for analysis • The method used to copy these regions is the polymerase chain reaction or PCR A close look at PCR • • Manipulation, Techniques, amplifying, PCR amplification • Manipulation, Techniques, amplifying, making many copies, amplification graph Agarose gel electrophoresis to separate pieces of amplified DNA by size Which regions of the DNA are variable enough to serve as a fingerprint? • Not genes but the regions in between genes where there are repeat sequences. • Short tandemly repeated sequences are highly useful Animation of STR analysis • /d/index.html • Applications, Human Identification, profiling, Today’s DNA profile Uses of DNA profiling • Guilt or innocence in a crime • Paternity testing • Proved Bill Clinton had sex with Monica Lewinsky • Thomas Jefferson or a close relative fathered a son with Sally Hemings, his slave • Identify victims of mass causalties, like 9/11 Personalized Medicine. Hope or Hype? Diagnosing inherited forms of breast cancer: Up close and personal Up to 10% of breast cancer can be shown to be Denise- patient inherited and most of these cancers are traceable to mutations in the BRCA1 or BRCA2 genes. Genes and Medicine,Applications, gene testing, Denise, “Decision to have her breasts removed”. Dr. Barbara Weber- …Barbara Weber,”Seeing who has the marker”. clinician and scientist The science behind the medicine . Applications, Genes and Medicine, gene hunting, Markers animation. Applications, Genes and Medicine, gene testing, making a pedigree animation. Applications, Genes and Medicine, gene testing, Testing for BRCA1 and BRCA2 animation The end of the story Genes and Medicine,Applications, gene testing, Denise, “Was she at increased risk” and “Finding out”. What would you have done if you were Denise or a member of her family? Remember that this gene was passed down from a male ancestor and males have higher cancer rates if they carry these genes. New Frontiers: Personalized Medicine. Hope or Hype? Individual genome sequencing “Imagine the day when you and your doctor sit down to review a copy of your own personal genome. This vital information about your biology will enable your physician to inform you of your disease susceptibilities, the best ways to keep yourself healthy and how to avoid or lessen the impact of future illnesses.” Why whole genome sequencing? •Require full functional understanding. Government genome project was a man and a woman from Buffalo. Celera was Craig Venter’s genome. Am I the same as them? • So sequencing was selective and missed: -personal mutations -regulatory mutations in noncoding sequence -repetitive sequences because they are difficult to sequence The phase problem? Does the linear arrangement along the length of a chromosome have an impact in addition to the genotype? Is the upper arrangement “better” than the lower one? Are there “cis” effects in the arrangement of the genes on chromosomes? “Personal DNA” Neil DeGrasse Tyson Nova Science Now copy copy of the 2010 commencement speech youtube fragment within the whole PBS program The Promises of Personalized Medicine •Provide advanced screening for disease. •Select safer, more effective medications and dosages. •Generate better vaccines. Develop vaccines made of genetic material. These would activate immune system w/o causing infection. Inexpensive, stable, easy to store. Engineered to carry several strains of a pathogen at once. Develop cell-based vaccines. •Lower healthcare costs. Proactive lifestyle changes, early detection & treatment. Decrease adverse drug reactions. Decrease # medications, length of time on medication. Are We There Yet? Reality Check. •Field of personalized medicine is in its infancy. •Access to personal genome is a long way off , or maybe not. $5000 genome by 2009 – Complete Genomics •For personalized medicine to be realized will require that we greatly reduce the cost and increase the speed of human genome sequencing. •This is the goal of the Archon X PRIZE for Genomics. Personalized Medicine: Genomics and Stem Cell therapies. Genomics: Example: Cancers – Tumor genotyping 2009 Science 326, 218-220. lung cancer non-small cell lung cancer most common EML4-ALK fusion others: mutations in EGFR, Iressa Genotype every lung cancer. Check for array of mutations. Target drug therapy based on specific genes mutated 15-20% of patient tumors can be matched to specific drug. Reality: better initially, then cancers develop resistance. NCI: The Cancer Genome Atlas 20 cancer types, 5 years, $275 million in first 2 years. Disease T argets for Intervention. •ALS – Amyotrophic Lateral Sclerosis •Huntington’s Disease •Type I Diabetes •Cancers •High Blood Pressure •Mental Illnesses •Asthma •Many are the same diseases that are targeted for stem cell therapies. • Cell therapies also referred to as regenerative medicine. NIH Stem Cell Information: A video stem cell primer Video Stem Cells Neil deGrasse Tyson Nova Science Now copy What are stem cells and why are they important? Three important characteristics: 1. Capable of dividing to renew themselves for long periods. proliferation through mitosis long-term self-renewal 2. Unspecialized or undifferentiated. 3. Can be induced to differentiate. blood, heart muscle, nerve, etc. BUT, what are the signals that prevent differentiation during the self-renewal divisions? This is a key question! University of Wisconsin There are two main types of stem cells. 1. Embryonic stem cells. derived from embryos in vitro fertilized eggs 2. Adult stem cells. population in every tissue typically generate same type of differentiated cells. University of Wisconsin originally thought NOT to be pluripotent Recent experiments have revealed -hematopoietic stem cells of bone marrow can be induced to form neurons and heart muscle. -liver stem cells induced to produce insulin. How Are Human Embryonic Stem Cells Obtained? •Cleavage-stage embryo produced by in vitro fertilization. •~5 days to reach blastocyst stage. •Identify specific surface markers found only on undifferentiated ES cells. These surface markers change once cells begin to differentiate. In suspension culture, hES cells differentiate. multicellular aggregate ⇩ gelatinized solid medium ⇩ differentiate further 2007: Shinya Yamanaka’s lab & James Thomson’s lab Genes for reprogramming differentiated cells to iPS cells. •Using retroviruses as carriers 4 human genes were identified that could cause a differentiated cell to be reprogrammed as an induced pluripotent cell (Oct3/4, Sox2 with Klf4 and c-Myc) when introduced simultaneously with the virus. •Oct3/4 & Sox2: Master transcriptional regulators. •Klf4 & c-Myc are required to increase induction efficiency. HOWEVER, Viral vectors increase the risk of tumorigenicity. c-Myc is an oncogene. ~20% mice develop tumors. S. Yamanaka. 2009. Cell 137, 13-17.d_pluripotent_stem_cells 2008: Generation of Induced Pluripotent Stem Cells without viral vectors. Shinya Yamanaka’s lab: single plasmid Expresses c-Myc plus cDNAs for Oct3/4, Sox3, Klf4 Transfection in mice. iPS cells without plasmid integration. Produced teratomas when transplanted into mice. Produced adult chimeras. Used embryonic fibroblasts. 2008 Science 322, 949-53 Generation and application of iPS Cells S. Yamanaka 2009 Movies Neil Theise CBS January 23, 2009 local copy Where America Stands: Stem Cell Research CBS March 16, 2010 local copy Potential Uses of Human Stem Cells •Spinal injuries and neuronal regeneration. •Skin cell replacement •Cardiovascular disease •Insulin producing pancreatic cells You have had the science, but what are the ethical issues for hES or iPS cells? •consent & confidentiality of donors coded but anonymous •exploitation of young women •animal-human chimeras reaching adults integration of human genes into nonhuman animal tissues nonhuman primates of particular concern no animal with hES cells introduced should be allowed to breed •religious concerns •other ethical issues Attendance exercise: For or against stem cell research and therapy? Post to Bulletin Board


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