Chapter 6.1 + 6.2 Introduction to Biotech and DNA tools used in biotech ✓ Flashcards
(25 cards)
Define biotechnology
Biotechnology refers to the artificial tools used on organisms or the products of organisms to make a product or solve a problem for human benefit.
What are the 4 DNA biotechnology techniques?
-DNA sequencing
-Polymerase chain reaction (PCR)
-Gel electrophoresis
-Recombinant DNA technology
What are the four tools used in DNA-based biotechnology?
-Restriction enzymes
-DNA ligase
-DNA polymerase
-Primers
What are these 4 DNA-based tool required for?
They are required for synthesising, cutting and pasting, viewing and analysing DNA.
What is the role of restriction enzymes?
-Restriction enzymes role is to cut DNA molecules at recognition sites (specific nucleotides) usually 4-5 bases long.
What are the two types of cuts done by restriction enzymes?
-Sticky ends (one strand has overhanging complementary bases, specific)
-Blunt ends (no overhanging, non-specific)
What is the role of DNA ligase?
-It is an enzyme that uses formed phosphodiester bonds to seal and reassemble the DNA backbone in the process of ligation, or in short, it sticks the backbone of DNA together.
What is the main source of restriction enzymes?
Bacteria
What is the role of DNA polymerase?
It is a class of enzymes that synthesises new strands of DNA based on a template strand and according to complementary base-pair rules.
What is the role of primers?
They assist in the synthesis of new strands of DNA by acting as a signal for the polymerase to begin synthesising.
What are primers?
Primers are short fragments of single-stranded nucleic acid (DNA or RNA)
What are the advantages of both sticky end and blunt end?
Sticky end: Fragments can join efficiently with a desired fragment that is cut with the same restriction enzyme.
Blunt end: Fragments can join with any other blunt end fragment
What enzyme synthesises primers?
Primase
Which end of DNA strands do primers stick to?
The 3’ end of each strand
Define recognition site
A specific sequence of DNA at which restriction enzymes will cut
Recall the bond that forms when DNA ligase joins two fragments of DNA
phosphodiester (sugar-phosphate) bond.
Contrast sticky-end and blunt-end cuts made by restriction enzymes
Sticky-end cuts:
-Cut DNA asymmetrically
-Leave overhanging single-stranded ends
-Overhangs can base pair with complementary sequences
-Easier to join with matching DNA fragments
Blunt-end cuts:
-Cut DNA straight across both strands
-No overhanging bases (ends are flush)
-Cannot base pair naturally
-Harder to join without special enzymes
-(cannot easily stick to other DNA fragments on their own because they have no overhanging bases to match with another strand)
Explain why primers are needed when DNA is being synthesised?
-DNA polymerase cannot start a new DNA strand on its own; it can only add nucleotides to an existing strand.
-Primers provide this starting point.
-In cells, primers are short RNA sequences made by primase, while in PCR, they are short synthetic DNA sequences that bind to the template strand, allowing DNA polymerase to begin synthesis
If a restriction enzyme made one cut in a plasmid and one cut in a piece of linear DNA, explain why the number of fragments that result differs?
A plasmid is circular, so one cut opens the circle into a single linear fragment.
Linear DNA already has two ends, so one cut splits it into two separate fragments.
What are restriction endonucleases?
Also known as restriction enzyme, is an enzyme that cuts DNA at specific restriction sites
What are restriction fragments?
A short fragment of DNA generated when a restriction enzyme cuts a longer DNA sequence
What is DNA sequencing?
The process of establishing the nucleotide sequence of a piece of DNA
What is polymerase chain reaction?
A cyclic method used to rapidly amplify (replicate) relatively small amounts of DNA into large amounts for further laboratory uses such as gel electrophoresis and DNA profiling
What is gel electrophoresis?
A technique that separates large molecules (either fragments of DNA or proteins) according to their size and charge for visualisation and identification purposes
