Gene Technology- Genome Projects and Making DNA Fragments Flashcards
What is a genome?
The entire set of DNA, including all the genes in an organism
What have improvements in technology allowed us to do?
To sequence the genomes of a variety of organisms
What do gene sequencing methods only work on?
Fragments of DNA which are sequenced and then put back in order to give the sequence of the whole genome
What was the Human Genome Project and when was it completed?
Mapped the entire sequence of the human genome for the first time in 2003
What is a proteome?
All the proteins that are made by an organism
What type of organisms don’t have much non-coding DNA?
Simple organisms such as bacteria
Why is it helpful when organisms don’t have much non-coding DNA?
It is relatively easy to determine their proteome from the DNA sequence of their genome
When can it be useful for organisms not having much non-coding DNA?
In medical research and development
Why is it harder to translate the genome of complex organisms?
They contain large sections of non-coding DNA and they also contain complex regulatory genes (determine when the genes that code for particular proteins should be switched on or off)
Why does containing large sections on non-coding DNA make it more difficult to translate the genome of complex organisms into their proteome?
Because it’s hard to find the bits that code for proteins among the non-coding and regulatory DNA
What were sequencing methods like in the past?
Labour-intensive, expensive and could only be done on a small scale
What are sequencing methods like now?
Automated, cost-effective and can be done on a large scale
What does recombinant DNA technology involve?
Transferring a fragment of DNA from one organism to another
What are organisms that contain transferred DNA known as?
Transgenic organisms
What are the three ways that DNA fragments can be produced?
Using reverse transcriptase, using restriction endonuclease enzymes and by using a gene machine
Why is it difficult to obtain a DNA fragment containing the target gene?
Most cells only contain two copies of each gene
Why is mRNA easier to obtain?
Cells contain many mRNA molecules which are complementary to the gene
How can mRNA molecules be used?
As templates to make lots of DNA
What does reverse transcriptase do?
Makes DNA from an RNA template
What is the DNA called that is produced from an RNA template?
cDNA- complementary DNA
What do pancreatic cells have?
Loads of mRNA molecules complementary to the insulin gene, but only two copies of the gene itself
What can reverse transcriptase be used for in pancreatic cells?
Making cDNA from the insulin mRNA
How does reverse transcriptase make cDNA from the insulin mRNA?
- mRNA is isolated from cells
- Mixed with free DNA nucleotides and reverse transcriptase
- The reverse transcriptase uses the mRNA as a template to synthesise a new strand of cDNA
What are palindromic sequences of nucleotides?
Sequences that consist of antiparallel base pairs
What are restriction endonucleases?
Enzymes that recognise specific palindromic sequences and cut the DNA at these places
Why do different restriction endonucleases cut at different specific recognition sequences?
Because the shape of the recognition sequence is complementary to the enzyme’s active site
When can you use restriction endonucleases to separate the fragment from the rest of the DNA?
If recognition sequences are present at either side of the DNA fragment you want
How is the restriction endonuclease used?
The DNA sample is incubated with the specific restriction endonuclease which cuts the DNA fragment out via a hydrolysis reaction
What can restriction endonucleases leave?
Sticky ends
What are sticky ends?
Small tails of unpaired bases at each end of the fragment
What can sticky ends be used for?
To bind (anneal) the DNA fragment to another piece of DNA that has sticky ends with complementary sequences
How has technology recently been developed?
So that fragments of DNA can be synthesised from scratch without the need for a pre-existing DNA template
What does the database involved in gene machine contain?
The necessary information to produce the DNA fragment
How does the gene machine work?
- Sequence required is designed if it doesn’t already exist
- First nucleotide in the sequence is fixed to some sort of support
- Nucleotides are added step by step in the correct order, in a cycle of processes the includes adding protecting groups
- Oligonucleotides are produced
- Broken off from the support and all the protecting groups are removed
- Oligonucleotides can the be joined together to make longer DNA fragments
What do protecting groups do?
Make sure that nucleotides are joined at the right points, to prevent unwanted branching
What are oligonucleotides?
Short sections of DNA
