January 07, (adenine, guanine, cytosine, thymine)

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1 (adenine, guanine, cytosine, thymine)

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3 DNA at Work - DNA is used to make proteins - proteins are made by linking amino acids (there are 20 possible amino acids) - sequence of amino acids determines shape/function of protein - each group of three bases on DNA codes for an amino acid DNA Replication - semi-conservative process (one of the original strands is maintained in the new strand) - DNA strand unwinds, and exposed bases are matched up by complimentary base pairing - requires many enzymes - DNA polymerases are enzymes that assemble new DNA in the proper base sequence determined by the original strand

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5 DNA Replication in Forensics - DNA polymerases are used to copy a DNA strand outside a living cell - called polymerase chain reaction (PCR) - small amounts of DNA can be copied with the aid of DNA polymerase - uses a machine called a DNA Thermal Cycler Restriction Enzymes: - chemicals that act as scissors to cut DNA molecules at specific locations - used in recombinant DNA - a portion of DNA is cut out with the aid of restriction enzymes - that portion is then inserted into a foreign DNA strand (usually from bacteria) - the new DNA is then reintroduced into the bacterial cells and the altered DNA is copied and passed on

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7 Restriction Fragment Length Polymorphisms (RFLP) - not all nucleotide sequences in DNA code for proteins - tandem repeats are sequences of letters repeated numerous times that do not code for a protein - help forensic scientists distinguish one individual from another - all humans have the same type of repeats, but there is variation in the number of repeats we each have RFLP: - repeats that are cut out of DNA by a restriction enzyme - forensic scientists use a number of different RFLPs for DNA typing - sequence is bases long and repeats itself up to 1000x

8 RFLP Typing - restriction enzymes cut up chromosomes into hundreds of fragments - some fragments contain repeating sequences from DNA - this creates different fragment lengths - it is these fragment lengths that allow forensic scientists to distinguish one person from another - once DNA is cut up by restriction enzyme, resulting fragments are sorted out by electrophoresis - cut up DNA is placed in separate lanes on electrophoresis gel and subjected to an electric field which causes them to move - smaller fragments move faster than larger ones - once electrophoresis process is over, DNA fragments are chemically treated so that the strands separate from each other - fragments are then transferred to a nylon membrane (Southern blotting) and treated with radioactive probes and X- rayed

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14 Polymerase Chain Reaction (PCR) - a technique designed to copy or multiply DNA strands - use DNA polymerase - to do PCR, sequences of DNA on each side of the region to be multiplied are identified - DNA strands are heated, so double helix separates - primers are added to separated strands and they combine with the DNA strands - DNA polymerase is added and so are free nucleotides - the polymerase rebuilds the DNA double helix - this doubles the number of DNA strands - this is repeated until you have ample copies of the DNA - RFLP DNA strands are much longer than strands generated by PCR - PCR can copy less than 1 billionth of a gram of DNA

15 Short Tandem Repeats (STR) - latest method of DNA typing - STRs are locations on the chromosome that contain short segments of three to seven repeating base pairs - one commonly used STR is TH01 - this DNA segment contains repeating sequence A-A-T-G - during a forensic examination, TH01 is extracted from biological materials and copied using PCR - the copies are then separated on electrophoretic gel - by examining the distance the STR migrates on the gel, the number of A-A-T-G repeats can be determined - every person has two STR types for THH01 - one inherited from each parent - hundreds of STRs are found in the human genome

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17 Capillary Electrophoresis: - preferred technology for the characterization of STRs - carried out in a thin glass column rather than on a gel plate - each end of capillary is immersed in a buffer that holds electrodes to supply high-voltage energy - DNA containing sample solution is injected into one end of the column - STR fragments move through the column bc of the electric potential at a speed related to the length of the fragment - other end of column is attached to a detector that tracks the separated STRs as they emerge from the column - it is recorded on an electropherogram

18 Electropherogram Y-axis is relative fluorescence (rfu) X-axis is size of DNA

19 Mitochondrial DNA (mtdna) - found in the mitochondria - inherited solely from the mother - constructed in a circular configuration - mtdna can be often found after nuclear DNA has degraded - can find an abundance of mtdna in skeletal remains - forensic analysis of mtdna is more rigorous, time consuming and costly than nuclear DNA profiling - used mtdna to identify the remains of the soldier in the Tomb of the Unknown Soldier

20 Combined DNA Index System (CODIS) - all fifty states have legislatively mandated the collection of DNA samples from convicted offenders of particular crimes and have established DNA databases for law enforcement purposes - CODIS was developed by the FBI and maintains local, state, and national databases of DNA profiles from convicted offenders, unsolved crime-scene evidence, and profiles of missing people - enables the comparison of DNA profiles electronically - US has standardized on thirteen STRs for entry into CODIS

21 Can get DNA from: - baseball bats/weapons (sweat, skin, blood, tissue) - hat, bandanna, mask (sweat, hair, dandruff) - eyeglasses (sweat, skin) - facial tissue/cotton swab (mucus, blood, sweat, semen, ear wax) - dirty laundry (blood, sweat, semen) - toothpick (saliva) - cigarette butt (saliva) - stamp or envelope (saliva) - tape or ligature (skin, sweat) - bottle, can, glass (saliva, sweat) - used condom - inside and outside (semen, vaginal/rectal cells) - blanket, pillow, sheet (sweat, hair, semen, urine, saliva) - bullet (blood, tissue) - bite mark (saliva) - contact lens (eye cells)

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