Genome Projects And Gene Technologies Flashcards
Genome:
The entire set of DNA in an organism.
How can we sequence the genomes of a variety of organisms?
Chopping up DNA into smaller fragments.
Proteome:
All the proteins made by an organism
Why is it easier to determine the proteome of bacteria from their genome?
Because they don’t have much non-coding DNA.
What is Recombinant DNA Technology?
It involves transferring a fragment of DNA from one organism to another.
How can we make DNA fragments using Reverse Transcriptase? (2)
- mRNa molecules are used as a template to make lots of DNA using an RNA template - produces cDNA.
- to do this, mRNa is isolated from cells and mixed with DNA nucleotides and reverse transcriptase.
How can we synthesis fragments of DNA from scratch? (2)
- gene machine
The sequence that is required is designed and the first nucleotide is fixed to some sort of support
Nucleotides are added step by step in correct order
In the gene machine, projecting groups are added, what are they?
They make sure that the nucleotides are joined at the right points to prevent unwanted branching.
What does the gene machine produce?
Short sections of DNA called oligonucleotides - they’re joined to make longer DNA fragments.
What aré restriction endonucleases / enzymes?
Enzymes that recognise specific palindromic sequences (complementary bases) and cut the DNA at these places.
Why do different restriction enzymes cut at specific recognition sequences?
Because the shape of the recognition sequence is complementary to the enzyme’s active site.
So overall what happens:
- DNA sample is incubated wirh the restriction enzyme which cure the DNA fragment out via a hydrolysis reaction.
Sometimes restriction enzymes cut and leave “sticky ends” what is this?
Small tails of unpaired bases at each end of the fragment - they can be used to bind the DNA fragment to another piece of DNA with complementary sequences sticky end.
What does in vivo amplification do?
Make lots of copy of the DNA fragment using a vector DNA.
What is a vector DNA?
Something that’s used to transfer DNA into a cell e.g. plasmids.
Describe how the DNA fragment is inserted into a vector: (3)
What is produced?
- DNA fragment inserted into vector DNA.
- vector DNA is cut open using the same restriction enzyme used to isolate it.
- vector DNA and DNA fragment are mixed with DNA ligase which joins the sticky ends of the DNA fragment to vector DNA - ligation.
- produces recombinant DNA.
How does the vector transfer the DNA fragments into the host cells: (4)
- vector w recombinant DNA is used to transfer the gene into host cells.
- if a plasmid vector is used, host cells need to be persuaded to take in the plasmid vector and DNA.
- with a bacteriophage vector, the bacteriophage will infect the host bacterium by injecting its DNA into it - the phage DNA then integrated into the bacterial DNA.
- host cells that take up the vectors are said to be transformed
How can we identify the transformed host cells? (3)
- market genes inserted to vector at the same time as gene being clones.
- host cells are grown on Agar plates. Transformed cells produce colonies where all the cells contain cloned and marker gene.
- marker gene can code for antibiotic resistance so only the transformed cells will survive and grow.
What would you do if you wanted the transformed host cells to produce the protein coded for by the DNA fragment?
The vector needs to contain specific promoter and terminator regions.
What are promotor regions?
DNA sequences that tell the enzyme RNA polymerase when to start producing mRNA.
