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DNA is the hereditary material

Experiments:

Griffith: mouse and pneumonia


-this is just what it sounds like
-Griffith got two strains of pneumococcus bacteria which infect mice – one non-
virulent strain (rough) and one virulent strain (smooth)
-he injected the non-virulent strain (rough) into the mouse and the mouse
lived (duh)
-he injected the virulent strain (smooth) into the mouse and the mouse died
(duh)
-he injected heat-killed virulent strain (smooth) into mouse and mouse lived
-THEN he injected live non-virulent strain and DEAD virulent strain into
mouse…AND MOUSE DIED
-what does this mean? Means that the non-virulent had somehow been
“transformed” into the virulent strain by the “transforming principle” that was
somehow part of the dead virulent strain (smooth) we now know that DNA is the
“transforming principle”

Avery
-basically a follow-up to Griffith’s experiment – experimental demonstration that
DNA is the substance that causes bacterial transformation
-isolated active portion of virulent pneumococcus and tested it
-found that enzymes that break apart RNA and protein had no effect on it, but
enzyme that breaks apart DNA destroyed extract’s transforming power

Hershey-Chase: virus smoothie


-they were looking at a phage called T2, which infects E. coli
-they knew that T2, like most other phages, was composed almost entirely of
DNA and protein
-they also knew T2 phage could quickly turn E. coli cell into a T2-producing
factory that released many copies when the cell ruptured
-so they wanted to figure out whether DNA or protein was responsible for
this
-Experiment:
-they used a radioactive isotope of sulfur to tag protein in one batch of T2
-they used a radioactive isotope of phosphorus to tag DNA in a second batch
of T2
(this worked because sulfur largely concentrated in protein,
phosphorus in DNA)
-then they tested the two samples shortly after the onset of infection to see
which type of molecule (protein or DNA) had entered the bacterial molecules and
thus was capable of reprogramming they found that there was RADIOACTIVE
PHOSPHORUS in the bacterial molecules in the radioactive phosphorus samples but
no radioactive sulfur in the corresponding samples, which meant phage DNA entered
the host cells but phage protein did not!
-in addition, traces of radioactive phosphorus remained when the infection
ran its course
-so Hershey and Chase concluded that the DNA injected by the phage was the
molecule carrying the genetic info that made the cells produce new viral DNA and
proteins
-powerful evidence that nucleic acids are the hereditary material!

-double helix

-Franklin: crystal structure

-Chargaff: A=T, G=C

-Watson and Crick

-semi-conservative replication: Meselsohn and Stahl

Double Helix Basics

DNA replication
-replication of a DNA molecule starts at particular sites called origins of replication
– these are short stretches of DNA having a specific sequence of nucleotides
-proteins that initiate DNA replication recognize this sequence and attach to the
DNA, separating the two strands and opening up a replication bubble
-at the end of each replication bubble is a replication fork: Y-shaped region
where parental strands of DNA are being unwound
-several proteins participate in unwinding:
-helicases are enzymes that untwist the double helix at the replication forks,
separating the two parental strands and making them available as template strands
-after parental strands separate, single-strand binding proteins bind to the
unpaired DNA strands, keeping them from re-pairing
-the untwisting of the double helix causes tighter twisting and strain ahead of
the replication fork; topoisomerase helps relieve the strain by breaking, swiveling,
and rejoining DNA strands
-then, the unwound sections of parental DNA strands are available to serve as
templates for the synthesis of new complimentary DNA strands:
-the initial nucleotide chain produced during DNA synthesis is actually a
short stretch of RNA; the RNA chain is called a primer and is synthesized by the
enzyme primase
-primase starts a complementary RNA chain from a single RNA
nucleotide, adding RNA nucleotides one at a time, using the parental DNA strand as
a template; thus the completed primer (~5-10 nt long) is base-paired to the
template strand. New DNA strand will start from 3’ end of RNA primer
-DNA polymerases (enzymes) catalyze synthesis of new DNA by adding
nucleotides to a preexisting chain

ANTIPARALLEL ELONGATION (THIS IS SOME LEADING STRAND/LAGGING


STRAND SHIT)
-two strands of DNA are “antiparallel”, meaning they’re oriented in opposite
directions to each other
-so new strands formed during DNA replication have to end up antiparallel to their
template strands (this is talking about 5’3’)
-BECAUSE OF THEIR STRUCTURE, DNA POLYMERASES CAN ADD NUCLEOTIDES
ONLY TO THE FREE 3’ END OF A PRIMER OR GROWING DNA STRAND
-so a new DNA strand can elongate only in the 5’3’ direction
-leading strand: along one template strand, DNA polymerase (III) can
synthesize a complementary strand continuously by elongating the new DNA in the
mandatory 5’3’ direction; DNA polymerase remains in replication fork and
continuously adds nucleotides to the new complimentary strand as the fork
progresses. ONLY ONE PRIMER REQUIRED FOR DNA POL III TO SYNTHESIZE THE
LEADING STRAND
-lagging strand: to elongate the other new strand of DNA in mandatory
5’3’ direction, DNA polymerase has to work in the direction away from the
replication fork and so since DNA polymerase can only work in 5’3’ direction, it
has to work in small parts (Okazaki fragments) and there need to be RNA primers
for each Okazaki fragment

Polymerase Chain Reaction


-science tool for cloning DNA
-3-step cycle that brings about a chain reaction that produces an exponentially
growing population of identical DNA molecules
-1: denaturation: reaction mixture is heated to separate (“denature”) DNA
strands
-2: annealing: reaction mixture is cooled to allow annealing (hybridization)
of short, single-stranded DNA primers complementary to sequences on opposite
strands at each end of target segment
-3: extension: DNA polymerase extends the primers in the 5’3’ direction
-primers are CHOSEN so they hybridize (bond with?) ONLY with complementary
sequences at opposite ends of the target segment (and for specificity’s sake, they
should be at least ~15 nucleotides long)
-remember the little tree chain reaction picture for exponential growth
-
DNA REPAIR
-mismatch repair: enzymes remove and replace incorrectly paired nucleotides
resulting from replication errors
-nucleotide excision repair: nuclease (DNA-cutting enzyme) cuts out a strand
containing the damage and the resulting gap is then filled in with nucleotides using
the undamaged strand as a template (filling is done by DNA polymerase and DNA
ligase)

Chromatin
-complex of DNA and protein – tightly packed into the nucleus
-so there’s the double helix of DNA
-then you’ve got these proteins called histones, which are responsible for the first
level of DNA packing in chromatin
-nucleosomes – DNA + histone proteins

Telomeres

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