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For a molecule to serves as the genetic material, it must be able to:
- replicate
- store information
- express information
- allow variation by mutation
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Proteins were major candidate for genetic material becuase
- 1940s many geneticists favored proteins
- proteins are more diverse and abundant in cells
- subject of the most active areas of genetic research, with more known about proteins than nucleic acid
- DNA was thought of as too simple to be genetic material and only four types of nucleotides compared to 20 different amino acids
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Griffith (1927)
- showed that avirulent strains of Diplococcus pneumoniae could be transformed to virulence
- speculated that the transforming principle could be part of the polysaccharide capsule or a compound required for capsule synthesis
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Avery, MacLeod, and McCarty
demondstared that the transforming principle was DNA and not protein
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Hershey and Chase (1952)
- using E.coli and an infecting virus (bacteriophage T2) demonstrated DNA and not protein was genetic material
- used radioisotope 32P and 35S
- demonstrated that DNA enters the bacterial cell during infection and directs viral reproduction
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Transfection
- process of infection by viral DNA into bacterial cells
- proved conclusively that the viral DNA alone contains all the necessary information for production of mature viruses
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UV light
- DNA and RNA absorb UV light most strongly at 260 nm and UV light is capable of inducing mutations in genetic material at 260 nm
- This provided strong indirect evidence for DNA as genetic material
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Recombinant DNA technology
- strongest direct evidence for DNA as the genetic material
- segments of eukaryotic DNA corresponding to specific genes are isolated and spliced into the bacterial DNA
- complex can be inserted into a bacterial cell and monitored
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Tobacco mosaic virus
demonstrates that RNA serves as the genetic material for these viruses (some viruses have a RNA core instead of a DNA core)
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RNA replicase
replication of viral RNA
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Reverse transcriptase
- Retroviruses replicate in this unusual way
- RNA serves as template for synthesis of complementary DNA by the RNA dependent DNA polymerase
- DNA is incorporated into the host cell genome; when transcribe, copies of the original retroviral RNA chromosomes are also produced
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Nucleotides
- building blocks of DNA
- Consist of nitrogenous base, pentose sugar, and phosphate group
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Purines
- Adenine and Guanine
- Bicyclic compounds
- Both have an amine group
- Guanine has a carbonyl
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Pyrimidines
- Cytosine, Thymine and Uracil
- Cytosine = one amine group and carbonyl
- Uracil = two carbonyl groups
- Thymine = two carbonyls and a methyl group
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Nucleoside
- contains the nitrogenous base and the pentose sugar
- MISSING PHOSPHATE
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Nucleotide
nucleoside with a phosphate group added
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Where is the phosphate group located on the nucleotide?
Carbon 5
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The number of phosphate groups have what effect on nucleotides?
- their naming system
- ex. NMPs, NDPs, NTPs
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Phosphodiester bond
- this is how nucleotides are linked to each other
- they are linked at the carbon 5 and the OH group on the carbon 3 group
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Chargaff
- showed that the amount of A is proportional to T and the amount of C is proportional to G
- however percentage of CG is not necessarily equal to the percentage of AT
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X-ray diffraction
- studies by Rosalind Frankllin
- DNA showed 3.4 angstrom periodicity
- characteristic of helical structure
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Watson and Crick
- proposed that DNA is a right-handed double helix in which two strands are antiparallel and the bases are stacked
- two strands are connected by AT and GC pairing and there are 10 base pairs per helix turn
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How many hydrogen bonds are formed in AT base pairs?
two bonds
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How many hydrogen bonds are formed in GC base pairs?
three bonds
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A DNA
- slightly more compact than B DNA
- prevalent under high-salt or dehydration conditions
- doubtful that it occurs in vivo
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B DNA
- seen under aqueous, low-salt conditions and is believed to be the biologically significant conformation
- more compact that C DNA, D DNA, E DNA, and P DNA
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Z DNA
forms a left-handed double helix
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3 classes of cellular RNAs that function during expression of genetic information:
- messenger RNA (mRNA)
- ribosomal RNA (rRNA) makes up 80%
- transfer RNA (tRNA)
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rRNA
structural components of ribosomes for protein synthesis
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mRNA
template for protein synthesis
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tRNA
carry amino acids for protein synthesis
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Nucleic acid can be separated by
gradient centrifugation procedures
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Sedimentation equilibrium centrifugation
- separates by density gradient
- bouyancy
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Sedimentation velocity centrifugation
measures the velocity of sedimentation in Svedberg coefficient units
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Hyperchromic shift
- during DNA denaturation
- used to determine the melting temperature (Tm)
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Melting temperature
method for estimating the base composition of DNA
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Molecular hybridization
- denaturation and renaturation of nucleic acids
- increased understanding of gene transcription
- refinements have helped advance study of molecular evolution and organization of DNA in chromosomes
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Fluorescent in situ by hybridization (FISH)
- uses fluorescent probes to monitor hybridization
- can be used to identify the chromosomal location of a DNA of interest
- ex. DNA specific to the centromeres of human DNA
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Reassociation kinetics
- analyzes rate of reassociation of complementary single DNA strand
- provides information about he size and complexity of genomic DNA from an organism
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Nucleic acid electrophoresis
separates RNA and DNA fragments by size such that smaller fragments migrate through a gel at a faster rate than large fragments
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