Summary notes for AQA A-level Biology recombinant DNA topic. Includes clear information on recombinant DNA, plasmids, gene cloning (in vivo and in vitro), marker genes, PCR, DNA probes, DNA fingerprinting. Summarised from class notes and the official course textbook. From an A* student.
Producing DNA fragments
• A number of human diseases mean individuals can't produce certain chemicals e.g. insulin
• Previous treatment involved introducing the chemical from a human/animal donor however this
can lead to rejection, immune system issues and infection
• Techniques can now isolate genes, clone them and transfer them into micro-organisms which
are then grown to produce the required protein continuously
• The DNA of two different organisms that has been combined is called recombinant DNA and
the resulting organism is genetically modified/ transgenic
• Genetic engineering is the process of moving a gene from one species to another
Universal code
• Since DNA code is the same in all organisms then DNA can be transferred
• Mechanisms of transcription and translation are also very similar in all organisms
• This is indirect evidence for evolution
Process of making a protein:
1. Isolation of DNA fragments that have the
gene for the desired protein
2. Insertion of the DNA into a vector
3. Transformation - transfer of DNA into
suitable host cells
4. Identification of the host cells that have
successfully been transformed using gene
markers
5. Growth/cloning of the population of host
cells
Isolation of DNA fragments:
3 different methods:
1. Conversion of mRNA to cDNA using reverse transcriptase
2. Using restriction endonuclease to cut fragments containing the desired gene from DNA
3. Creating the gene in a gene machine based on a known protein structure
Using reverse transcriptase
• Reverse transcriptase catalyses the production of DNA from RNA
• A cell that readily produces the protein is selected
• The relevant mRNA is extracted
• Reverse transcriptase used to make DNA from RNA. The DNA is known as complementary DNA
(cDNA)
• To make the other stand of DNA, the enzyme DNA polymerase is used to build up the
complementary nucleotides
• These DNA fragments will be cloned in bacteria, and called a cDNA library
• Introns are not present in this method
, Using restriction endonuclease enzymes
• These enzymes are found in bacteria to defend themselves from viruses by cutting up the viral
DNA
• Each restriction endonuclease cuts a DNA double strand at a specific sequence of bases known
as the recognition sequence. It cuts at a specific point due to a specific active site that is
complementary to one base sequence.
• The recognition sequence is a palindromic sequence which means that the enzymes can cut
from either direction.
• Cuts in the DNA can form blunt ends or sticky ends
• Sticky ends: can forms hydrogen bonds with complementary sticky ends.
• To get complementary sticky ends, the same restriction endonuclease must be used
• Introns are present in this method
Fragments:
• This technique leaves you with many fragments, one containing the gene you want
• Smaller recognition sequences have a higher probability of occurring in the DNA so the
fragments will be shorter
• Different lengths of fragments from a different species due to the recognition sequences being
in different places as they have different genes
DNA types Plasmid Linear DNA
Picture
1 cut 1 fragment 2 fragments
2 cuts 2 fragments 3 fragments
Using a gene machine
• Desired sequence of bases determined from the protein’s primary structure and from this you
can deduce the mRNA codons and the complementary DNA triplets
• Desired sequence of nucleotide bases given to computer
• Computer designs small overlapping single strand of nucleotides (oligonucleotides)
• Each of the oligonucleotides are joined together to make a gene and this gene is replicated and
created into a double strand using the polymerase chain reaction
• Genes are checked and those with errors are rejected
The benefits of buying summaries with Stuvia:
Guaranteed quality through customer reviews
Stuvia customers have reviewed more than 700,000 summaries. This how you know that you are buying the best documents.
Quick and easy check-out
You can quickly pay through credit card or Stuvia-credit for the summaries. There is no membership needed.
Focus on what matters
Your fellow students write the study notes themselves, which is why the documents are always reliable and up-to-date. This ensures you quickly get to the core!
Frequently asked questions
What do I get when I buy this document?
You get a PDF, available immediately after your purchase. The purchased document is accessible anytime, anywhere and indefinitely through your profile.
Satisfaction guarantee: how does it work?
Our satisfaction guarantee ensures that you always find a study document that suits you well. You fill out a form, and our customer service team takes care of the rest.
Who am I buying these notes from?
Stuvia is a marketplace, so you are not buying this document from us, but from seller amybowler. Stuvia facilitates payment to the seller.
Will I be stuck with a subscription?
No, you only buy these notes for $3.89. You're not tied to anything after your purchase.
