GEN MICROBIOLOGY BIOL 2051
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This 9 page Class Notes was uploaded by Charles Kohler on Tuesday October 13, 2015. The Class Notes belongs to BIOL 2051 at Louisiana State University taught by K. Sullivan in Fall. Since its upload, it has received 8 views. For similar materials see /class/222793/biol-2051-louisiana-state-university in Biological Sciences at Louisiana State University.
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Date Created: 10/13/15
Selected topics from chapter 10 and 12 Chapter 10 Molecular Regulation Regulating Gene Expression Microbes respond to changing environment Alter growth rate Alter proteins produced Must w their environment Receptors on cellsurfi Must transmit information to chromosome Alter gene expression Change transcription rate copying DNA to RNA Change translation rate RNA making proteins make more proteins or less proteins Sensing the Environment Twocomponent signal transduction 1 Sensor kinase protein in m Binds to signal 0 Food 0 Chemicalcue 39 Activates itself 2 Cytoplasmic response regulator 39 Takes phosphate from sensor 39 Binds chromosome 0 Alters transcription rate of multiple genes Altering Transcription Rates sigma factor guides RNA polymerase to initiate transcription at promoter I Proteins can help guide sigma factor to promoter 39 Activator Binds to DNA sites next to promoter lncreases freguency of that gene s transcription I Proteins can M sigma factor from Mg 39 Repressor o Mto DNA sites next to promoter 0 Operator 0 Lower s freguency of that gene s transcription The amount of an enzyme in the cell can be controlled by decreasing repression or increasing induction the amount ofMthat encodes the enzyme For negative control the regulatory molecule is called a repressor protein and it functions by inhibiting mRNA synthesis The E coli lac Operon Lactose M is uses as nutrient source bu E coli 0 Cannot pass through cell membrane 0 Lactose permease allows entry 0 Disaccharide lactose must be cleaved into monosaccharides to be digested o Betagalactosidase lactose 0 People also make bgalactosidase o If not person is lactoseintolerant o The lacZ encodes bgalactosidase o The lac Yencodes lactose permease 0 Need both proteins to digest lactose Operon 0 Multiple genes transcribed from one promoter 0 Both genes are transcribed together Repressor protein Lacl transcription 0 Repressor binds to operator 0 Blocks sigma from bindingpromoter Repressor responds to presence of lactose 0 Bindsinducer allolactose or DNA not both 0 Add lactose gt repressor falls off operator From pic repressor is like a road block unless The lac operon is an example ofnegative control oftranscription Wilminisrml Transcription Marked Repressor ac erator i5 w I L p ara rmr WWW IacYVImml ImlW 39 gt Transcription prnzeeds Repressor b Inducer allolactose Forgositive control of transcription an activator protein binds to activator bindingsites on the DNAto simulate transcription 0 Fortranscription ofthe maltose operon 1 39 binds to the 39 39 proteins 2 maltose activator protein binds to the activator bindin site on the DNA RNA polymerase can then proceed with transcription The example of uan ulpllull Activator mal binding site Momma maIE muIF 11titrmnml WWW WWW 39 M WWW WWW 39 No transcription l Maltese activator protein a Activator Ina bindinte PromotermemalE WmalFm l maIG39mLy 1 Int W W MM Transcription 139 proceeds Maltese activator protein 1 lnducer maltose Catobolite Repression 0 Glucose is easiest sugarto digest catabolize o If glucose is present lac operon not transcribed other sugar operons 0 Presence of glucose af39fects signal inside the cell I Lowers amount of CAMP High glucose gt low CAMP 0 CRP protein is an activator protein 0 Binds next to promoter on the DNA 0 Increasestranscotpn oflacand otheroperons CRP responds to presence of CAMP o Acts as activator only when bound to CAMP 0 High glucose gt low CAMP levels gt CRP inactive Does not bind operons gt low level of Iactranscription The Iacoperon is underthe control of catabolte repression as well as its own specific egativ regulatog system Two requirements for induction of the lac operon 1 CAMP levels high enough for CPR protein to the CRPbinding site 2 lactose present The presence of glucose represses the lac operon by inhibition cAMP synthesis I CHaoter 10 psotive and negative something and something elese Chapter 12 Biotechnology Applied Microbial Biotechnology Use microbes to express genes 0 Produce eukaryotic proteins in large amounts I Human insulin Use microbial gene products 0 Microbes have huge diversity of enzymes I PCR depends on archaeal polymerase lmmunize against 0 Use wfrom pathogens I Without exposure to pathogenic organism lnsulin lnsulin is required for those with Type I diabetes amp many with Type II diabetes Prior to genetic engineering insulin was harvested amp purified from pig or cow pancreas Nonhuman insulin is less effective on humans some developed algeris to it 0 Human insulin was 151 human protein made commercially using engineered bacteria 0 ldentical in every way to human produced insulin 0 Today engineered yeast is used to make insulin Plant biotechnology o Glycophosphate Roundup herbicide resistant plants 0 Bacteria were found that were resistant to this herbicide o The gene for the herbicide resistance was put into some crop plants soybeans corn cotton Transgenic plants Insectresistant plants Bacillus thuringensis produces chemical Bttoxin that is toxic to larvae of moths butterflies beetles flies Gene for Bttoxin was put into cotton and potato plants 0 Expressed in leaves of plant o Kills only insects on those crops o Eliminates need to spray chemical pesticides onto crops I Chemicals are dangerous to use toxic to humans expensive Vaccines through Food Vaccines expose human to pathogens proteins 0 Expose immune system cells to proteins 0 We develop immune defenses to proteins 0 Protects us from later infection by pathogen 0 Proteins can be produced in plants absorbed via intestine o No longer need to grow pathogen to produce protein 0 Ex Bananas that have vaccine for cholera Gene therapy nonfucntianl gene is replaced by a functional gene Clone desired gene into viral genome a virus gets the functional gene that the human wants lnfect person with modified virus 0 Infected cells will express cloned gene I Make desired protein Genes for many diseases have been located and the mutations in the defective genes have been identified Huntingtons cystic fibrosis SCID Genetic engineering Making new DNA molecules Restriction enzymes each RE recognizes a specific DNA sequence and makes double stranded breaks at the sequence resulting DNA fragments can be separated using agarose gel electrophoresis DNA fragments from different DNA molecules can be joined together using ligase to make recombinant DNA PCR Polymerase chain reaction method used to make many copies of a specific DNA sequence in vitr0outside of the body using primers and DNA polymerase First described in 1985 Nobel Prize for Kary Mullis in 1993 made possible by discovery of Taq polymerase DNA polymerase of thermophile from hot spring Thermus aquaticus materialsreagents used in PCR DNA nucleotide building blocks for the new DNA Template DNA the DNA sequence that you want to amplify Copy Primerss single stranded DNAs between 20 and 50 nucleotides long oligonucleotides that are complementary to region on either side of the temple DNA DNA polymerase a heat stable enzyme that catalyzes the synthesis of new DNA 3 major steps in PCR repeated for 20 to 40 cycles in automated Thermal Cycler which can heat and cool reaction tubes quickly Denatureation Step N 94 C double stranded DNA melts to single stranded DNA Annealing N 54 C Primers hydrogen bond to complementary sequences of DNA template Extension N 72 C Deoxyribonucletides complementary to template are added to the primer on the 339 end In theory you can start with one and end up with a lot which is 39 httpwwwsumanasinccomwebcontentanimationscontentpcrhtml httpwwwdnalcor ddnalcresourcesshockwavepcranwholehtm oo lecom videosearchhlenamprlscom microsoften quS H5Dwampresnum1ampqPCR20animationampum1ampieUTF 8ampsaNamptabwvqPCRamphlenampembO Iron nun Agarose gel electrophoresis Used to determine size of DNA fragments DNA samples are loaded into slots at one end of an agarose gel An electrical current is passed through the gel DNA has negative charge so moves toward the positive electrode smaller DNA fragments move farther than the larger fragements Using a size standard size of erach band is knownsize of other fragments can be determined Negative electrode SM FgtQquotPU39l0l l quotIZMWNGQ Positive electrode httpwwwsumanasinccomwebcontentanimationscontentgelelectrophoresishtml there are a couple of test questions that come from this bacteria cover us help us educate our immune systems Vibrio fishcheri makes light comes from the ocean whats interesting is when the bacteria makes light when they come to a certain cell number they all light up so how do they do this they talk to each other when alone no light but make ans secret harmones but when there are a lot of bacterias they secrete the molecule and they all turn all the light live in hawaiia squid make light squid wants light uses light from bacteria to anitillumite light so it uses it to not get a shadow and be eaten The bacteria doesn t stay in the bacteria all the time just at night when needed All bacteria can talk to each other called Bacteria always control themselves with So if one bacteria gets in you and secrests hormones nothing happens but if a bunch of baceteria get together and secrete hormone at the same time they can infect you Something allows bacteria to count their own siblings Bacteria not only talk to themselever but they are multilingual have a second enzyme that makes a second signal language in interspecies communication how many of me and how many f you and think about what task to take out depending on the populations How to kill bacteria anitbitoics are running out so we stop bacteria from talking to each other and from senesing the populations of them and us interspecies communication are targeted 9 how When bacertium gets into animal doesn t intiatew virulence reacgnies when engouh for aatack animal dies so what do we do Is we treat animal with a pathogenic bacterium and antisomething bacteria
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