DNA FingerprintingEach DNA profile is Unique Alec Jeffreys, 1985
ContentsSome basic factsIntroductionHistory of DNA fingerprintingStages of DNA fingerprintingMethods of DNA fingerprintingApplications •Crime •Food •Miscellaneous
Some baSic factS . When the number is On DNA sequence of not known, nucleotide variable, or Is repeated againirrelevant, itis sometimes and again but differ called for person to person a variable number When between 10 and 60 nucleotides are tandem repeated – Mini- satellite repeats (VNTR) Those with fewer are known as microsatellites or short tandem repeat
introduction It isthe number of times that these STRs arerepeated that produces Regions of chromosomes that code for Extrons contain blocks of repeatedthe variations proteins are called introns. nucleotides short tandem repeats in (STRs) individuals. Other regions that are non-coding are called extrons.
introduction . DNA fingerprinting on the basis of STR doesn’t give an Individual DNA fingerprint but able tofingerprint determine the two ing samples are form the Is also same person called "DNA fingerprinting" is a technique that DNA allows us to see differences in DNA profiling fragments from one person to another in more general form.
HiStory of dna fingerprinting•Up to 1984, the only method ofestablishing and authenticatingpersonal identification was by thefingerprint process.•DNA fingerprinting technique wasdevised in 1985 by Alec Jeffrey atUniversity of Leicester in England,while working on the sequenceswithin myoglobin gene.
Stages of DNA Fingeprinting DNA is extracted from sample•Each cell is surrounded bycell membrane•DNA is enclosed within thenucleus
Stages of DNA Fingeprinting DNA is extracted from sampleTo extract DNA , most commonly used detergent is SodiumDodecyl Sulfate (SDS). Digestio n of proteins and lipids around the cell
Stages of DNA Fingerprinting DNA is extracted from sample•DNA in thenucleus of the cell ismolded, folded, andprotected by Proteins•Proteinase K isused to break up theproteins
Stages of DNA Fingerprinting Ethanol PrecipitationThe DNA must be purified from the cell extract.Ethanol lowers the effective water concentration, causinglarge bio-molecules to interpenetrate and aggregate. The resultis a visible precipitate at the interface, where the ethanol isconcentrated. DNA will rise into the alcohol layer from the cell extractlayer. The protein and grease parts dissolve in the bottom, waterylayer
Stages of DNA FingerprintingDNA cuts with the help of RestrictionEndonucleaseDNA is cut into millions of small fragments with the help ofrestriction endonucleasesRestriction enzymes chop DNA at specific sequences.the patterns occur in different places in different individual , thelength of the fragment differs from person to person.e.g The EcoRI restriction enzyme recognizes the following sequence -C-A-A-T-T-G- -G-T-T-A-A-C
Stages of DNA FingerprintingSeparation of DNA fragments•DNA fragments are separatedby electrophoresis•DNA is negatively chargedso it is attracted to the positiveend of the gel. Separated DNA on the basis of gel electrophoresis
Stages of DNA Fingerprinting Southern blotting•Patterns of fragmentsare transferred to a nylonmembrane by a process calledSouthern blotting.
Stages of DNA Fingerprinting DNA fingerprints comparednylon or nitrocellulose (Fromsouthern blot) sheet is placed underX-ray filmThe radioactive probes on theDNA fragments expose the filmproduces visible pattern of lightand dark bands which is unique toeach individual Light and dark bands of DNA when nylon film exposed to X- ray
Methods of DNA Fingerprinting Restriction enzyme based fingerprinting•Relatively straight forward•When genomic DNA is cleaved with particular restriction enzymes, theresulting fragments can contain a minisatellite (VNTR) region•gel electrophoresis separates the digested DNA into bands based on thelength of the fragments, the pattern of bands reflects the number ofrepeats in a minisatellite•each person’s DNA will yield a unique banding pattern
Methods of DNA Fingerprinting PCR – based DNA fingerprinting• By virtue of the amplification process, it is possible to begin with onlya tiny amount of DNA• possible to fingerprint the DNA of a single cell• Also possible to analyze old or degraded samples using PCR-basedfingerprinting• Degraded DNA - be nearly impossible to fingerprint with restrictionanalysis
Methods of DNA Fingerprinting Amplified fragment Length PolymorphismCome in 90’sremains attractive because of its relatively less complicated operationand the cost-effectivenessuse of gel in its analysis phase, there are issues of bunching of theVTRNs, causing misidentifications in the process.
Methods of DNA Fingerprinting STR ( Short tandem repeats)Mostly widely used method for comparing samples of personsSTR analyzes how many times base pairs repeat themselves on aparticular location on a strand of DNAHas been extremely successful for use in the personal identification ofcriminal suspects, paternity issues, as well as in identification of thediseases.
Applications of DNA fingerprinting Mostly used to solve crime and medical, paternity problems •Violent murder occurred •Rape cases • Inheritance cases •Immigration cases•DNA fingerprinting also found recent applications in foodindustry•Recent application in breast cancer detection
Applications of DNA Fingerprinting Crime scene• Pattern of theDNA profile iscompared with thoseof the victim and thesuspect• profile matches thesuspect - providesstrong evidence thatthe suspect waspresent at crime scene
Applications of DNA fingerprinting Application in FoodProduct identificationGenetic diversity analysesTraceability testing
Applications of DNA fingerprinting Product identification.Based on found genetic similarities or differences, aproducts identity can be established.makes it possible to distinguish raw materials orproducts from different suppliersHigh-value products, such as Basmati rice andArabica coffee that cannot always be easily andreliably distinguished from inferior varieties.
Applications of DNA fingerprinting Genetic diversity analysisDNA fingerprints - been applied in the genetic diversityanalysis of wine cultivarssome varieties consisted of more than one genetically distinctvariety, and that varieties that had been considered distinct weregenetically identicalSuch an analysis is interesting for many crops and forms thebasis for the analysis of variations in product quality.
Applications of DNA fingerprinting Traceability testingDNA fingerprinting technology can be used for traceabilitytestingcomparison of a fingerprint of a cows ear and the fingerprintof meat products will show whether the two samples originatefrom the same animal
Applications of DNA fingerprinting Commercial sauerkraut fermentations Process available generates more salt as waste – disposableproblemTo avoid this, one has to study bacterial ecology – previous itwas studied by bio- chemical methodsBut , DNA fingerprinting found application – shows morecomplex behavior of bacterial viruses which produces waste
Applications of DNA fingerprinting Commercial sauerkraut fermentations so , on the basis of these data, one can generate newfermentation technology – which produces less salt waste andincreases the yield
Applications of DNA fingerprinting Wines – use of GM yeast and GM grapes Scientists at the Bavarian State Institute for Viticultureand Horticulture in Würzburg, Germany, use DNAfingerprinting to analyze wines.Genetically modified (GM) yeast and GM vines areundergoing field trials
Applications of DNA fingerprinting False labeling issue Wang Jun and his group at the Chinese University of HongKong used DNA fingerprinting to differentiate between Panaxginseng (ginseng) and Panax quinquefolius (American ginseng)price of P. quinquefolius is usually 5–10 times greater thanthat of cultivated Panax ginseng
Applications of DNA fingerprinting Accessions ( distinct varieties) of plants accessions are traditionally classified on the basis ofmorphology, but it is suspected that numerous duplicates arepresentproblem is increased by the fact that the same accessions areoften named differently in different countries
Applications of DNA fingerprinting Accessions ( distinct varieties) of plants twenty-one accessions of sweet potato from the SouthAfrican genebank were compared.RAPD and SSR techniques were usedResults show that 4 accessions which are differently namedare the genetically same – DNA fingerprinting one can use toidentify and make gene banks of world more appropriate
Applications of DNA fingerprinting Lactic acid bacteria DNA fingerprints of lactic acid bacteria were generated byPCRmethod made it possible to identify 37 isolates from raw milk,industrial starters and yogurtDifferentiation at species, subspecies and strain level waspossible for Lactobacillusdelbrueckii subsp. lactis, Lb.delbrueckii subsp bulgaricus and Str.thermophilus
Applications of DNA fingerprinting Miscellaneous• Fingerprinting has provided them to determine whether awoman who had breast cancer at about as low as 10 cancercells In evolutionary studies, DNA fingerprinting has eventraced the origin of all of mankind to Africa•fingerprinting can also be used to trace plants and plantproducts back to their sources
concluSion .One can use DNA fingerprinting isDNA very useful technique tofingerprinting solve many problems related to crime, rapeIn near era of and murder cases whenfood science to very few evidences arewiden and availableknow more •DNA samples must be prepared carefullyabout theproducts •Also , STR based method of DNA fingerprinting is very easy and less time consuming