DNA Tech & Its Application Flashcards
What is the recombinant organism?
Bacteria’s plasmid+ foreign DNA= plasmid put back into the bacteria——> recombinant bacterium
What is the restriction site and restriction enzyme?
Restriction site= interest gene on the DNA, restriction
endonuclease cut,
When the restriction enzymes cut the sequences, how was the DNA in the restriction site protected?
By adding methy groups to adenine or cytosine.
What’s PCR( polymerase chain reaction)
Amplification of the gene by making copies of the genes of a specific sequence.
Includes 3 steps- denaturation or heating to separate DNA strands
Annihilation for primers and complementary hydrogen bondings at 3’ ends.
Heat stable DNA polymerase to replicate DNA.
How many DNA strands does one cycle of PCR produce?
Formula for the number of DNA strands after each cycle= 2 power n
After one cycle-2
After 2 cycle - 4
Is PCR used in cell cloning?
No, although PCR is rapid and specific, it induces lots of error and hence corrected sequences or good copies that can be used are limited.
What does PCR also contain to be able to do gene cloning?
Restriction sites at the end of DNA fragments like those in plasmids DNA for restriction enzymes to cut and bind together with other DNA to induce recombinant DNA.
If eukaryotic genes are put into prokaryotic cells like bacteria, what needs to be done?
To overcome the differences in prokaryotes and eukaryotic DNA control sequences, expression vectors, a highly active bacterial promoter should be introduced at the upstream of the restriction site. So that bacteria can recognize it and express foreign genes.
Since bacteria cannot excise introns, what are the consequences?
Incorrect expression. Therefore, sequences that contain exons should only recombined.
What eukaryotic organism have plasmids like bac?
Yeasts- can also do splicing and post translational modifications for the expressed proteins.
What’s in situ hybridization?
Scientists want to know the location of the cloned genes or genes that are producing cancer cells in the body.
They produce nucleic acid probes ( DNA probes or RNA probes). Dye it with fluorescent and the probe hybridize with the complementary base pairing on the mRNA sequences. We can know the exact position of the hybridization place= in situ hybridization
What is RT-PCR and why is it used?
To access the genes of differential developmental stages.
MRNA—> cDNA—> RT- PCR- electrophoresis
What is the gel used for electrophoresis?
What energy is used to perform?
Agarose
Electric current passes through it
What is microarrays?
DNA chips, containing many different genes+ cDNA probe+ fluorescent
What is RNA sequencing?
Actually cDNA sequencing- cDNA sequenced with computers, results containing the number of times a sequence is present, indicate which genes are expressed in a given tissue at what level