Topic 1 - SACE Biology Stage 2

Nucleotide

a monomer of nucleic acids; contains a pentose sugar, a phosphate group, and a nitrogenous base

Nucleoside

nitrogenous base + sugar

deoxyribose

A five-carbon sugar present in DNA

ribose

A five-carbon sugar present in RNA

complementary base pairing

In DNA, T pairs with A; G pairs with C;

RNA, U pairs with A and G pairs with C

semi-conservative replication

in each new DNA double helix, one strand is from the original molecule, and one strand is new

helicase

An enzyme that untwists the double helix of DNA at the replication forks.

DNA polymerase

Enzyme involved in DNA replication that joins individual nucleotides to produce a DNA molecule

Hydrogen bonds

Type of bonding found between complementary bases

3

The number of hydrogen bonds found between the complementary nitrogenous bases G and C

2

The number of hydrogen bonds found between the complementary nitrogenous bases A and T

histone

Protein around which DNA wraps in eukaryotic cells in chromosomes

gene locus

specific location of a gene on a chromosome

chromatin

DNA in its normal uncondensed state in the nucleus.

gene

A segment of DNA on a chromosome that codes for a single functional protein or a single polypeptide or RNA molecule

chromosome

an independent section of the entire DNA genome that appears as a threadlike structure carrying genetic information in the form of genes

strand of DNA

single string of nucleotides connected to each other via the sugar phosphate backbone

5' to 3'

direction DNA is read

molecule of DNA

two strands of DNA joined together by complementary base pairing between nitrogenous bases

single circular chromosome

DNA is found in this form in prokaryotic (bacterial) cells

nucleus and mitochondria

DNA is found in these two locations in eukaryotic cells

nucleic acid

macromolecule made of a chain of nucleotides eg DNA, RNA

nucleosome

Bead-like structure in eukaryotic chromatin, composed of a short length of DNA wrapped around a core of 8 histone proteins

endonuclease

another name for a restriction enzyme

restriction enzyme

Enzyme that cuts DNA at a specific sequence of nucleotides (4-8 nucleotides)

sticky ends

Single stranded ends of DNA left after cutting with restriction enzymes

primer

short sequence of single stranded DNA that is complementary in sequence to the start and end of a sequence of DNA to be copied

5' to 3'

Direction DNA is read

Oligonucleotide

a short polymer of nucleotides (usually fewer than 50) eg 10-30. Often used as primers.

Taq polymerase

Heat stable DNA polymerase from heat resistant bacteria Thermus aquaticus used in PCR

short tandem repeats

STR stands for......

non-coding

STRs are found in the........................regions of DNA

Negative

DNA has a ..............................charge due to the presence of phosphate groups

positively

DNA moves towards the .........................charged end of the polyacrylamide gel during gel electrophoresis

polyacrylamide gel

type of gel used to separate DNA fragments in gel electrophoresis

smallest

The...................DNA fragments travel the furthest along the gel in gel electrophoresis

Dideoxynucleotide

nucleotide that has hydroxyl groups removed which is used during DNA sequencing to terminate synthesis by blocking DNA polymerase

gene probe

Fluorescently or radioactively tagged DNA or RNA sequence that is complementary in sequence to a target DNA sequence. Used to locate the DNA sequence.

Transgenesis

transfer of genetic material from one species to another

recombinant DNA

DNA produced by combining DNA from different sources

bacterial plasmids

a tiny ring of DNA in the cytoplasm of bacteria that can replicate independently

recombinant plasmid

Plasmid that has an extra gene from another source inserted into it

Cluster Regularly Interspaced Short Palindromic Repeat

CRISPR stands for..........

gRNA (guideRNA)

Sequence of RNA used in CRISPR that is complementary in sequence to the target gene (DNA). Part of the Cas9 complex which guides where Cas9 cuts the DNA.

Cut DNA at a specific nucleotide sequence

Function of the enzyme in the Cas9 complex

guideRNA and enzyme

Cas9 complex consists of......

thermal cycler

An instrument that automatically cycles through different temperatures used to complete PCR reactions

polymerase chain reaction (PCR)

A technique for amplifying DNA in vitro by incubating with special primers, DNA polymerase molecules, and nucleotides.

forward, reverse

Two types of primers required in PCR

Denaturation

First stage of PCR that involves heating mixture to 95-98˚C to break hydrogen bonds and separate DNA strands

Annealing

Second stage of PCR that involves cooling mixture to 56˚C so complementary DNA primers bind to the DNA

Polymerisation

Final stage of PCR that involves heating mixture to 72˚C to activate the Taq polymerase so free nucleotides bind by complementary base pairing to form the new complementary DNA strand

doubles

The number of copies of DNA ................. with every cycle of PCR

Probe or dye

Possible methods used to visualise DNA fragments on gel electrophoresis

capillary electrophoresis

A method of separating DNA samples based on the rate of movement of each component through a gel-filled capillary while under the influence of an electric field

Sanger method of DNA sequencing

A method used to determine the nucleotide sequence of a piece of DNA by selective incorporation of chain-terminating dideoxynucleotides by DNA polymerase during in vitro DNA replication

DNA replication stops

Effect of tagged dideoxynucleotides (dATP, dGTP etc) on DNA replication when using the Sanger method

Electropherogram

data output from capillary electrophoresis where fluorescent signals are recorded as graphical peaks

Introns can be removed

Advantage of using recombinant yeast plasmids instead of recombinant bacterial plasmids

micro-injection

foreign DNA is inserted directly into a cell's nucleus using a glass micropipette

electroporation

Electrical current forms pores in cell membranes

homology directed repair

a mechanism in cells to repair double strand DNA breaks that relies on using the homologous chromosome as a repair template. This mechanism is used in genome engineering to incorporate precise genetic changes.

Non-homologous end joining

A mechanism for repairing double-strand breaks in DNA that involves quickly bringing together, trimming, and rejoining the two broken ends; results in a loss of information at the site of repair.

CRISPR-associated proteins

Cas stands for ..........

Vectors

Bacterial and yeast plasmids and viruses can be used as ......................... for transgenesis

Longest

The first peak seen on an electropherogram corresponds to the shortest/longest DNA fragment

DNA ligase

an enzyme that joins the sugar-phosphate backbones of DNA strands back together

Metabolism

◦ All chemical processes within an organism that sustain life

Anabolism

◦ The "building up" reactions

Eg synthesising DNA from free nucleotides

Eg photosynthesis

Catabolism

◦ The "breaking down" reactions

◦ Eg digesting proteins to amino acids

lower activation energy

Impact of enzymes on activation energy of reactions

Enzymes

Biological catalysts for chemical reactions in living organisms

protein

Enzymes are made of ................................

Substrate

reactant in an enzyme-catalysed reaction

active site

a region on an enzyme that binds to a substrate (protein or other substance) during a reaction

Conformational change

Term given to the slight change in shape to the enzyme's active site when substrate binds

Induced Fit Model

Model which explains that the enzyme active site undergoes conformational change as substrate binds

cofactor, temperature, pH

Factors that Effect Enzyme activity

cofactor

An inorganic compound or ion that binds to the enzyme (not the active site) which causes the active site to change shape and better bind the substrate. The enzyme is activated.

denaturation

loss of normal shape of a protein caused by heat or other factor due to breaking of bonds between localised portions of the polypeptide chain

Denatured protein

◦When a proteins secondary, tertiary, or

quaternary structure is disrupted

◦ Peptide bonds are not broken

◦ Often irreversible

primary structure/peptide bond

These bonds are not broken when proteins are denatured

secondary, tertiary, or quaternary

These levels of structure are disrupted when proteins are denatured

primary structure

Protein folding is determined by the ________________

competitive inhibitors

molecules that compete with the substrate for binding at the active site

non-competitive inhibitors

Bind to an allosteric site (not the active site) and deform the enzyme and its active site (conformational change), stop the substrate from binding indirectly

non-competitive inhibitors

In the presence of a lot of substrate, which type of inhibitor would be most effective?

allosteric site

A specific receptor site on some part of an enzyme molecule remote from the active site.

allosteric inhibition

the mechanism for inhibiting enzyme action in which a regulatory molecule binds to a second site (not the active site) and initiates a conformation change in the active site, preventing binding with the substrate

enzyme

Biological catalyst that speeds up a specific chemical reaction in a living system but is not used up in the reaction

activation energy

the minimum amount of energy required to start a chemical reaction

active site

a region on an enzyme that is complementary in shape to a substrate or substrates

substrate

reactant in an enzyme-catalysed reaction

anabolic

any chemical reaction that builds up complex molecules from simpler molecules

catabolic reaction

any chemical reaction that breaks down complex molecules into simpler molecules

induced fit hypothesis

Theory of enzyme catalysis which states that the partial binding of a substrate to an enzyme alters the structure of the enzyme so that its active site becomes complementary to the structure of the substrate, enabling tighter binding

competitive inhibitor

Molecule that competes with the substrate for the enzyme active site

non-competitive inhibitor

a molecule that binds to an enzyme at a location outside the active site and inhibits the enzyme's function

slows down

effect of presence of competitive and/or non-competitive inhibitors on enzyme catalysed reaction rate

speeds up

effect of presence of co-factors on enzyme catalysed reaction rate