1.7 Biotechnology tools and techniques

DNA Extraction

Genetic material can be manipulated by scientists who transfer DNA from one species to the DNA of another

mRNA codons code for the same amino acid in every organism

Method

1.      Obtain cell/tissue sample

2.      Centrifuge the cells at high speeds to separate the nucleus from other organelles

3.      Add detergents to physically breakdown the nuclear membrane, exposing the chromosomes

4.      DNA Is isolated from histone proteins

Gene probes

Single stranded DNA or RNA segments that have been constructed in the laboratory

Have radioactive or fluorescent markers that allow them to readily locate genes

Once bound to the probe the gene is said to have been hybridised with the probe

Step 1: target DNA strands must be separated

Step 2: mixed with probe that complementary base pairs to the gene of interest

Step 3: cooled and detected using UV lights

Restriction enzymes

Bacteria produce restriction enzymes which cut DNA naturally and act as a defence mechanism against viral DNA

Used in genetic engineering to cut DNA at specific sites to remove or isolate genes

DNA ligase is used to join the host and donor DNA- bacterial transformation

Polymerase chain reaction (PCR)

Can be used to create multiple copies of the same DNA for analysis

Ingredients:

Target DNA template DNA must be copied

DNA primers – stops target DNA strands from reforming, binding site for Taq DNA polymerase

Free DNA nucleotides

Taq DNA polymerase – can tolerate higher temperatures than human polymerase

Buffer solutions

Step 1:  Denaturation – template DNA is heated so DNA bonds are broken

Step 2: Annealing- mixture is cooled so primers can bond to complementary bases

Step 3: Extension- nucleotides bind to exposed DNA bases and Taq DNA polymerase joins them into a strand

Gel Electrophoresis

Technique used to separate parts of DNA according to their size

DNA samples are loaded into wells (negatively charged electrodes)

As DNA is negatively charged the samples will move towards the positively charged electrons

The larger the fragment the slower it moves

DNA profiling

Technique that identifies the unique genetic make up of individuals or compare DNA from different sources

Short tandem repeats are sections of DNA (2-6 base pairs) that are repeated a number of times

DNA sample is collected

Restriction enzymes digest the sample

PCR is used to amplify the STRs

Gel electrophoresis is run

DNA sequencing

More accurate form of DNA analysis

Determines the sequence of bases in a gene segment or the entire genome

How it works

DNA template is mixed with Taq DNA polymerase, free floating DNA nucleotides, sequencing primes and terminator nucleotides

DNA is amplified by PCR, when a terminator nucleotide is added by Taq DNA polymerase, DNA synthesis is stopped

Electrophoresis is used to order fragments

Results displayed in an electropherogram