2.7 DNA replication, transcription and translation Flashcards
How do we know DNA is semi conservative and how can we prove it
semi-conservative DNA means that each of teh original strands serves as a guide or template for a new strand. It uses complementary base pairing of nucleotides. The new strand is tehrefore identical to the previous strand.
This was worked out through the Stahl and Mendelson experiment where they used nucleotides that had an rare isotope of nitrogen that is lighter. The DNA was seperated using density gradient centrifugation. The fact that a band for the denser N remained shows that the nucleotides are not broken up but semi conserved.
How is DNA replicated
The DNA unzips and DNA helicase breaks the H bonds of the DNA and toposiomerase keeps the strands apart.
The RNA primer attaches to the beginning of the unravelled strand. It attaches with RNA primase whihc gudies teh DNA polymerase to where it needs to add complementary polynucleiotides.
Complementary nucleotides attack to the DNA strands and the DNA Polymyerase 3 binds the new hydrogen bonded nucleotides together, forming the phosphodiester bon between the nucleotides forming the polynucleotide chain.
DNA polymerase 3 con only form the phosphodiester bond in teh 5’–> 3’ direction. The strand in the 5’ to 3’ end is the leading strand and is continuously formed. The strand in the other direction, forms okazaki fragments and is the lagging strand. Each with a RNA primer.
DNA polymerase 1 removes the RNA primer nucleotides allowing DNA nucleotides to come in and fill the gaps.
DNA ligase tehn joins the gaps left from the primer and seals the remaining DNA nucleotides together.
what is teh PCR and how is it carried out
a technique used to make mnay copies of a selected DNA sequence.
- double stranded DNA is separted into single strands by heating it so the H bonds break
- an excess of primer is added and the solution is cooled
- DNA polymerase is then used to add nucleotides to the strand and when enough time has elapsed the process repeats
what is transciption
the synthesis of RNA using DNA as a template,
- enzyme polymerase binds to start of gene
- rna polymerase moves along the gene separting dna into single strands and pairing RNA nucleotides
- rna polymerase forms covalent bonds between rna nucleotides
- rna separates from DNA and double helix reforms
- transcriptions stops at the end of the gene and the complete RNA molecule is released
The DNA wiht the same base sequence as the rna is the sense strand and the DNA strand that is transcribed is the antisense srtrand.
what is translation
this is the synthesis of a polypeptides on ribosomes
takes place in the cyotplasm on ribosomes which have binding sites for each of the molecules that take part in the translation.
- mRNA binds to the small subunit of hte ribosome
- a molecule of tRNA with an anticodon complementary to the first codon to be translated on teh mRNA binds to the ribosome
- 2 of these tRNA can bind at one time
- the ribosome transfers the amino acid carried by teh tRNA to teh 2nd tRNA via a peptide bond
- the second amino acid then becomes the frist and the tRNA is released
- the process continues until teh stop codon is reached,
the production of human insulin in bacteria
previously used porcine (pig) and bovine (cattle) insulin which has one and three amino acid differences respectively and bind to human insulin receptors
however can become allergic to this so use genetically modified e coli cells
all use same genetic code as humans so this works
Transcription of protein synthesis
mRNA and rRNA is opened up into cistrons wihc are sections of 17 bases that unravel - this means teh H bonds break between bases and the histrones break off
the 5-3 strand is the sense strand and teh 3-5 prime is the antisense strand which acts as a template in transcription
ythe enzyme RNA polymerase bind to a site on the DNA at the start of a gene, moves along the gense separating dna and pairing up RNA molecules, it forms covalent bonds between RNA nucleotides, it separtes from dNA and the helix reforms
activation of DNA
this is post transcriptional modification
1 - a cap is added to teh 5’ end which is a guanine this signals that the mRNA is ready to be translated
2 - a tail is added to the 3’ end which is 100 adenine, this prevents the mRNA from being broken down in teh cytoplasm by enzymes
3 - the RNA has exons and introns (coding and non coding regions) and splicing takes place so the introns are cut out and the extrons are fused together
teh nuclear envelop has tiny pores and the mRNA coils up and is then pushed out into the cytoplasm where they will attach to the ribosome
differences in protein synthesis in prokaryotes and eukaryotes
in prokaryotes there is no nucleus so there is less hindrance as it occurs in the cytoplasm - the nucleoid region
teh DNA that is transcribed is in the from of euchromatin so it is a littel bit unravelled in G1 as it needs to be loose enough to transcribe so the enzymes can get in to DNA
translation
mRNA gets attached to polysomes (the small subunit of a ribosomes) (many ribosomes at once) and it translated simultaneously so as to produce many copies of that protein
in translation teh mRNA is read in anticodon arrangement
- non overlapping fashion of reading read in 3s
- genetic code is universal
- genetic code is degenerate, 64 possible codons but only 20 possible amino acids
at the end of translation the poly a tail will come away
a tRNA with complementaryanticodons binds to the ribosomes and a second complementatry anticodon binds besides it
the ribosome transfer the amino acid from teh frst oto the second and forms a peptide bond, a dipeptide is created,
the tRNAs move up a spot and the first tRNA is removed, a new tRNA joins and the process is repeated
the production of human insulin in bacteria
