DNA Replication Part 1

Sp@BIO
DNA Replication process
INTRODUCTION
Replication can broadly be defined as genome duplication process in which a template derived nucleic acid synthesis take place when the template and the growing strands both are DNA.The unit of DNA in which an individual act of replication occurs is called replicon or replication unit. It includes that part of the DNA containing an origin of replication subtended by termination sequences on its both sides. In case of prokaryotes there is a single replication unit, whereas in eukaryotes, there are many replication units or replication.
MODELS OF DNA REPLICATION
There are three model of DNA replication -
i) Conservative Model - According to this, the two parental strands in the double helix remain completely conserved directing the synthesis of progeny double helix whose both the strands are synthesized new. This model does not fit good with the concept of DNA strand complementarity and hence rejected.
ii) Dispersive Model - This model was proposed by Max Delbruck after his theory of breakage and reunion, according to which segments of parental and progeny strands are interspersed as a result of the synthesis and rejoining of short segments of DNA. This concept also turned out to be incorrect.
iii) Semiconservative Model - This was first proposed by Watson and Crick (1953), according to which each single strand of a double helix acts as a template for the synthesis of new progeny strand. Thus in the daughter chromosomes one strand is old i.e parental and the other strand is new i.e. progeny strand. This means the half of the parental DNA remains conserved and its counter half part is newly formed. This semiconservative mode of replication is the natural mode of replication that exists in all living cells and can also be demonstrated in vitro.
EVIDENCE OF SEMICONSERVATIVE REPLICATION
Evidence in Prokaryotes (Messelson and Stahl’s experiment) -
The first critical test that DNA replication occurs by Semiconservative mechanism was performed by Mathew Meselson and Franklin Stahl in 1958. Their results show that, the chromosome of common colon bacteriaE.Coli replicate semiconservatively.
Meselson and Stahl took cells that had been grown for several generations in medium containing 15NH4Cl as nitrogen source. Due to this all the DNA become incorporated with 15N containing nucleotides. After growing for a number of generations, the cells were then removed from 15N medium, washed and transferred to 14N containing medium. Since 15N is the heavier isotope than its normal counterpart14N, the DNA of 15N cells will have a greater density than the DNA of14N cells. After the cells were allowed to grow in presence of 14N for varying periods of time (or generations), the DNAs were extracted and analyzed in CsCl2 equilibrium density gradient centrifugation. Their results as has been depicted in the figure- were found consistent only with the semiconservative mode of replication.
DNA isolated from cells grown for several generations in 15N medium, designated here as 0th generation cells, exhibited maximum density and formed band near the bottom of the centrifuge tube. This is regarded as heavy DNA band or H band. When DNA of1st generation 14N cells were subjected to ultracentrifugation the band was lifted up to a position of medium density. This DNA band was designated as hybrid DNA band or Hy. At this stage conservative mode could be said to be failed because if it had been true two bands were expected. The heavy DNA band (H) would have existed and a new band of lighter DNA would have been formed. Absence of H band in 14N grown 1st generation cells clearly rejects the possibility of conservative model. But whether the shift of band from high density region to medium density region occurred due to semiconservative replication or due to dispersive mode of replication could not be distinguished after 1st generation. But this result is of semiconservative replication became clear only after the 2nd generation of14Ngrown cells. After 2nd generation there were two bands formed in the centrifuge tube. The hybrid band remained as it is and a newer band of lighter DNA appeared. The inference drawn from these results revealed that this could only be an outcome of semiconservative mode of replication. It was further proved by the fact that the light band became wider and wider in consecutive generations (results not shown). Had it been dispersive mechanism, there would have been a gradual upward shift of band in each generation and in each generation only a single band lighter than the previous generation would have appeared. Therefore, these results were precisely the semiconservative mode of replication identical to the one predicted by Watson & Crick.
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[@debabratalaik9296]

I heard this topic description from you in tuition...so learning video...Thank you so much for giving another chance to hear it. #Regards

[@DrSanjeevPandey]

Thank you Raj...