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The image depicts the 3 main steps of OE-PCR Reaction. Besides the introduction of mutations, Overlap Extension PCR is widely used to assemble complex DNA sequences without the introduction of undesired nucleotides at any position.
Eukaryotes initiate DNA replication at multiple points in the chromosome, so replication forks meet and terminate at many points in the chromosome. Because eukaryotes have linear chromosomes, DNA replication is unable to reach the very end of the chromosomes. Due to this problem, DNA is lost in each replication cycle from the end of the chromosome.
During DNA replication, the replisome will unwind the parental duplex DNA into a two single-stranded DNA template replication fork in a 5' to 3' direction. The leading strand is the template strand that is being replicated in the same direction as the movement of the replication fork.
DNA is read in the 3' → 5' direction, therefore, nucleotides are synthesized (or attached to the template strand) in the 5' → 3' direction. However, one of the parent strands of DNA is 3' → 5' while the other is 5' → 3'. To solve this, replication occurs in opposite directions.
A large part of DNA (more than 98% for humans) is non-coding, meaning that these sections do not serve as patterns for protein sequences. The two strands of DNA run in opposite directions to each other and are thus antiparallel. Attached to each sugar is one of four types of nucleobases (or bases).
Replication of DNA always begins at an origin of replication. In yeast, the origins contain autonomously replicating sequences (ARS), distributed throughout the chromosome about 30 kb from each other. They allow replication of DNA wherever they are placed. Each one is 100-200 bp long, and the A element is one of the most conserved stretches.
DNA from individual bacterial clones is sequenced and the sequence is assembled by using overlapping DNA regions. Large-scale sequencing often aims at sequencing very long DNA pieces, such as whole chromosomes, although large-scale sequencing can also be used to generate very large numbers of short sequences, such as found in phage display.
Control of the DNA replication system ensures that the genome is replicated only once per cycle; over-replication induces DNA damage. Deregulation of DNA replication is a key factor in genomic instability during cancer development. [3] This highlights the specificity of DNA synthesis machinery in vivo. Various means exist to artificially ...