8 min readPathway sheetPart III (4 marks) or Part IV (7 marks): steps of recombinant DNA technology; the tools as 2-mark questions

Genetic engineering: recombinant DNA technology

Genetic engineering cuts out a useful gene with restriction enzymes and joins it into a vector with DNA ligase. A host cell then makes the product.

Genetic engineeringChanging an organism's genes in the laboratory by adding, removing or altering DNA

Restriction enzymesMolecular scissors: cut DNA at specific base sequences

DNA ligaseMolecular glue: joins pieces of DNA

VectorCarrier of the gene into the host: a plasmid or a bacteriophage

What is recombinant DNA?

Genetic engineering changes the genes of an organism in the laboratory. Recombinant DNA (rDNA) is DNA made by joining DNA from two different sources. An example is a human gene joined into a bacterial plasmid. The method of making it and putting it to use is recombinant DNA technology.

Tools of genetic engineering

Tool	What it does	Example
Restriction enzymes (restriction endonucleases)	Cut DNA at specific base sequences: molecular scissors	EcoRI, from the bacterium E. coli
DNA ligase	Joins the cut ends of DNA: molecular glue	Seals the gene into the plasmid
Vector	Carries the gene into the host cell	Plasmid (a small ring of DNA in bacteria, outside the main chromosome); bacteriophage (a virus that infects bacteria)
Host cell	Takes up the rDNA, multiplies and makes the product	E. coli bacterium, yeast

Steps of recombinant DNA technology

1
Isolate the DNA that has the desired gene, for example the human insulin gene.
2
Cut the DNA with a restriction enzyme to get the gene.
Restriction enzyme
3
Cut the plasmid with the same restriction enzyme, so the cut ends match ('sticky ends').
Restriction enzyme
4★
Join the gene into the plasmid. This new DNA is recombinant DNA.
DNA ligase
5
Insert the rDNA into a host bacterium such as E. coli. This entry is called transformation.
6
Multiply the host bacteria. Every copy carries the gene (gene cloning).
7★
Express the gene: the bacteria make the product, such as insulin, which is extracted and purified.

★ key step

Making human insulin in bacteria

How to draw the steps of rDNA technology

On the left, draw a long double-stranded DNA. Shade one part and label it desired gene. Draw small scissors at both ends of it and label restriction enzyme.
At the top middle, draw a small ring and label it plasmid (vector). Show a gap cut in the ring by the same enzyme.
Draw the gene fitting into the gap. Label DNA ligase and recombinant DNA.
Draw an oval bacterium with the ring inside it. Label host cell (E. coli) and transformation.
Draw several bacteria in a row coming from it. Label multiplication (gene cloning).
End with an arrow to product: human insulin. Write the title: Recombinant DNA technology.

In real life

Insulin for people with diabetes is now made mainly by genetically engineered bacteria or yeast. Earlier it was taken from the pancreas of cattle and pigs.

Exam angle

2 marks: “Why are restriction enzymes called molecular scissors?” They cut DNA at specific base sequences, like scissors, to remove a desired gene or to open a plasmid.

2 marks: “What is a plasmid? Why is it used as a vector?” A small circular DNA in bacteria, outside the main chromosome. It copies itself and can carry a foreign gene into the host.

7 marks: “Explain the steps of recombinant DNA technology with a diagram.” Tools (restriction enzyme, ligase, plasmid, E. coli); isolate gene, cut gene and plasmid with the same enzyme, join with ligase, transform host, multiply, express; human insulin; labelled diagram.

Remember it

Cut, paste, carry, copy: scissors (restriction enzyme), glue (ligase), vehicle (plasmid), factory (E. coli).