A2 Biology: Cloning

Cloning

• Genes cellsor whole organisms carrying identical genetic material
• Occurs naturally in
• identical twins
• plans which reproduce asexually
• bacteria replicating though binary fission

Asexual reproduction

• Prokaryotes divide by binary fission
• the DNA replicated and the cell divides
• Two cells are genetically identical 
• The basis of asexual reproductions in nearly all eukaryotes is mitosis
• the genetic material replicated to separated to form two nuclei
• the nuclei contains exact copies of the original DNA
• the cells splits to produce two daughter cells that are cloned offspring
• Advantages
• quick - reproduces rapidly - take advantage of the environment
• alternative if sexual reproduction fails or is not possible
• all offspring have the genetic information to enable them to survive
• Disadvantage
• it does not produce genetic variation
• therefore if the environment changes or a new disease is introduce then they are all equally susceptible.

Artificial vegetative propagation 

• taking cuttings
• section of a stem is cut between the leaf joints
• end treated with plant hormones
• plnted = forms ne plants
• grafting
• shoots section is joined to an already growing root and stem
• the graft grows and is genetically identical to the parent plant, but rootstock is genetically different
• advantages
• plants with useful features are identified for growth
• some have to be grown this way - bananas have to be cloned because cultivaed bananas are sterile
• farmers know what features the plant will have - yield, taste, disease resistance etc
• cost reduced because all the crop can be harvested at the same time
• faster
• disadvantages
• plants equally susceptible to new pests or disease
• crops are grown in specific areas and are divided by a certain distance to limit this effect

CLONING ANIMALS

• a cloned animal is one that has been produced using the same genetic information as another animal
• such an animal has the same genotype as the donor organism
• nuclear transfer - dolly the sheep

• splitting embryos
• an embryo can be genetically manipulated
• IVF using the sperm and egg of high value ales and females
• +ve
• high value animals produce in large numbers
• rare animals cloned and preserved
• genetically modified animals can be quickly reproduced
• -ve
• high value animals not produced with animal welfare in mind
• genetic uniformity reduced ability to cope with changes in the environment
• it is unknown how the process effect the long term health of the animals

Non-reproductive cloning

• this is the use of cloned cells to regenerate cells, tissue and organs to replace those that become damages
• +ve
• no rejection by the immune syste
• cloning and cell culture could end the wait for transplantation
• cloned cells are totipotent - generate any cell type (some diseses cannot be treated by transplant)
• likely to be less dangerous
• possibilities include:
• regeneration of heart tissues after a heart attack
• repair or nervous tissues destroyed by diseases such as multiple scleorosis
• repairing the spinal cord
• 
  

A2 Biology: Biotechnology

Biotechnology involved the exploitation of living organisms or biological processes to improve agriculture, animal husbandry, food science, medicine and industry

Food production

• Yoghurts and cheese
• produced by bacterial growth in milk
• this chnages the texture and flavour
• the bacteria prevent the growth of those bacteria that caue spoilage
• this helps preserve ood
• 'Qourn'
• produced by the growth of a fungus 'Fusarium'
• The fungal mass (mycelium) is processed and shaped into food
• Soy Sauce
• Roasted soya beans are fermented with yeast of fungus such as Aspergillus.

Drugs and pharmaceuticals

• Penicillin
• this is a product of the growth of a fungus called Penicillium.
• It is a by-product of the organisms metabolism
• this is isolated from the growth medium and is used as an antibiotic
• Insulin
• Bacteria such a E.coli are genetically modified to carry human insulin gene
• they secrete insulin as they grow

Bioremediation of waste products

• a variety of bacteria and fungi use organic waste in the water as nutrients and make wast harmless
• e.g. Fusarium grown on corn steep liquor, a waste product of the corn milling industry.

Why use bacteria and Fungi?

• grows rapidly in favourable conditions
• can produce chemicals or proteins that are released into the surrounding medium and ce be harvested
• can be genetically modified to produce specific products
• grow well at relatively low temperatures
• can be grown anywhere in the world - not climate dependant
• generated products that are in a pure form than those generated via chemical processes
• can often be grown using nutrient materials that would otherwise useless or even toxic to humans

GROWTH CURVE

What is a culture?

• it's a growth of micro-organisms
• it may be pure culture (single species) or mixed culture (mix of species)
• they can be cultured in liquid broth or solid nutrient agar gel.

Lag phase - organisms adjusting to conditions, cells active but not reproducing. Population is fairly constant

Log phase - population size doubles each generation. plenty of space and nutrients to reproduce.

Stationery phase - nutrients level decrease, waste products builds up. Death rate = production rate

Death phase - nutrients exhaustion, increased level of toxic waste products. Death rate > reproduction rate.

FERMENTATION

• Fermentation refers to the culturing of micoorganisms
• they could grow both aerobically and anaerobically in tanks called fermenters

Fermenters

• growth of an microorganism on an enormous scale
• begins with a 'start culture' a small pure culture from whicl all microorganis will grown
• conditions
• temperature
• type and tie of addition of nutrients
• O2 concentration
• pH

• Batch culture
• microorganisms mixed with specific quantity of nutrients then grown for a fixed period, no further nutrients added
• advantages and disadvantages
• growth rate slower
• easy to set up and maintain
• if contaminated, one batch is lost
• lless efficient - not operating all the time
• useful for producing secondary metabolites
• Continuous culture
• microorganisms mixed with nutrients and nutrients are added continuously  with products removed continuously throughout the process
• advantages and disadvantages
• growth rate higher
• difficult to set up and maintain
• if contaminated, huge volumes lost.
• ore efficient, operated continuously
• useful fro primary metabolites.
• Aseptic technique - asepsis
• unwanted micro-organisms can contaminate the fermentation process
• to avoid this, aseptic technique ensures contamination of the culture does not occur
• contamination may result in products being unsafe
• unwanted micro-organisms
• compete with culture micro-organisms for nutrients and space
• reduce the yield of the product
• may cause spoilage, produce toxic chemicals or destroy the culture micro-organisms

Metabolism and metabolites

• Metabolites include:
• chemicals such as hormones and enzymes
• waste products e.g. CO2, urea and O2
• these can be nutrients required by other organisms
• primary metbolites
• these are produced by organisms as part of its normal growth
• amino acids
• proteins
• enzymes
• ethanol
• lactate
• secondary metabolites
• these are produced by the organism and are not part of its normal growth
• antibiotic chemicals
• produced y a small number of microorganisms

Industrial enzymes

• Commercial use of enzymes
• Pectinase can be used in fruit juice extraction
• Metabolic enzymes in the bacteria A. niger produce citric acid used in detergents
• Downstreaming
• isolated enzymes can be produced in large quantities through biotechnology
• their extraction from the fermentation medium is known as downstream processing
• this is their separation and purification
• Immoilising enzymes
• when using enzymes in industry they need to be removed after they have served their purpose
• this can be a costly process
• it is possible to immobilise enzymes and prevent them from mixing
• immbolisation
• this is a technique were enzyme molecules are held, separated from the reaction mixture
• substrate molecules can bind to the enzymes and the products are formed go back into the reaction leaving the enzymes in place
• methods
• adsorption
• covalent bonding
• entrapment
• membrane separation 

A2 Biology: Roles of genes and environment in evolution

• The Hardy-Weingberg equation is used to show that population changes over time. (they evolve)

Environmental resistance: combined action of biotic and abiotic factors that limits the growth of a popuation.

Selection pressure: 

• environmental actor that confers greater chances of surviving and reproducing on some members of the population than others
  e.g. Rabbits that are well camouflaged are more likely to escape predation, survive, reproduce and pass on favourable (camouflaged) genes to offspring.
• Selection pressure in this case is predation
• it reduces the chances of white or black coat being passed on
• this keeps the population stable = stabilising selection
• If the environment changes - becomes colder, ore snow - rabbits with white fur would have selective advantage. White rabbits are more likely to survive, reproduce and pass white alleles to the next generation.
• frequency of white alleles would increase  = directional selection
• leads to evolutionary change.

Types of selection

• Driving force behind evolution is natural selection and this occurs in three main ways
• stabilising selection
• most common
• a response to a stable environment
• e.g. birth weight
• under weight babies are less likely to survive
• overweight babies are likely to get stuck during birth killing not only themselves but also their mothers.
• The result is that the population graph gets narrower and taller as selection agaisnt mutations takes place
• bottle neck event - occurs when a population is reduced to just a few breeding individuals (e.g. cheetahs)
• though the total population may later recover, they will all be descendants of the few originals and so will have a much smaller gene pool than the original population.
• directional selection
• results ina population of new trait
• takes place whenever a change occurs in the environment
• an evolutionary force of natual selection
• e.g. resistance. Wararin resistance in rats, DDT resistance in mosquitoes or antiiotic rsistance in bacteria. Resistant individuals soon become the dominant type within the population
• disruptive selection
• less common
• results in two distinct populations
• eventually these two forms may become so distinct they become new species

Isolating mechanism

• A large population of organisms may be split into sub groups that are prevented from interbreeding by
• geographical barriers e.g. rivers, mountain range
• seasonal/temporal barriers e.g. climate
• reproductive mechanism

Genetic drift

• change in allele frequency in a population, as some alleles pass to the next generation and some disappear
• large changes in small populations
• as the size of the population decreases the degree of fluctuation increases

SPECIES

Basic classifcation system

• Domain          Eukaryotes
• Kingdom        Animalia
• Phylum          Chordates
• Class              Vertebrata
• Order             Mammalia
• Family            Primates
• Genus            Great Apes
• Species           Home sapiens

Biological species concept

• a group of similar organisms that can interbreed and produce fertile offspring
• problems with this definition
• some species do not reproduce sexually
• some members of the same species look very different  (e.g. gender)

Phylogenetic species concept

• a group of organisms that have similar
• morphology
• physiology
• embryology
• behaviour
• and occupy the same ecological niche
• Closely related organisms will have similar DNA
• DNA analysis is carried out to compare particular base sequences (haplotype)
• Any differences  in bases is expressed a % divergence
• A clade is a group of all the organisms that share a single common ancestor and therefore have the similar DNa and features. 
• aka a monophyletic group e.g. A single ancestor and all its descendants

Cladistics

• based on evolutionary ancestry (phylogentic relationships)
• it does not use a fixed number of level such as kingdom, phylum and class
• whereas taxonomic classification focuses on similarities between organisms
• it include both monophyletic and paraphyletic groups

• Use molecule analysis e.g. DNA/RNA sequencing
• computer programs to represent evolutionary tree of life
• makes no distinction between extinct and existing species
• the cladisitic approcach has often confirmed the Linneasen classfication of orgnanisms but has sometime sled to organims being reclassified

Paraphyletic groups

• includes most recent ancestors but not all of its descendants e.g. reptiles
• Class: Reptilia. Reptiles are cold blooded vertebrates which unlike amphibians possess thick, impermeable skin covered y scaled. They have lungs, not gills and usually a three chambered heat. Their eggs are watertight shells. - Paraphyletic as it excludes birds which are descendants of reptiles
• The cl

A2 Biology: Variation

Interspecific variation - it's easy to see how one species of organism varies from another. e.g. elphant from a kangaroo.

Intraspecific variation - less obvious is the distinction between the two individuals of the same species but nevertheless there are small differences (thus no two elephants are found to be identical)

Cause of variation

• variation is the result of either genetic differences or the influence of the environment
• in many cases its a combination of both genetic and environmental factors

Genetic differences

• are due to different genes each individual possesses
• genetic variations arises as a result of
• mutations
• these sudden changes to genes and chromosomes may or may not be passed onto the next generations
• meiosis
• this special form of nuclear division forms the gametes - this mixes up the genetic material before it is passed on to the gametes all of which are therefore different.
• fusion of gametes
• in sexual reproduction the offspring inherit some characteristics of each parent and are therefore different from both of them
• which gametes fuses with which at fertilisation is a random process adding to the variety of the offspring.

Discontinuous variation

• Qualitative difference between phenotypes
• Some characteristics of organisms grade into one another; there are no separate categories but a complete range of values.
• examples
• ear lobe shapes
• ability to role tongue
• blood type
• gender
• due to
• influence of one or more genes
• if greater than one they behave epistatically
• if only one gene then its called monogenic
• different alleles at a single gene locus have large effects on the phenotype
• different gene loci have different effects on the phenotype
• e.g. codominance 

Continuous variation

• due to
• more than one gene - polygenic
• genes provide an additive effect to the phenotype
• different alleles at a gene locus have a small effect
• genes found on different chromosomes
• environmental influences
• affect the way the organisms' genes are expressed.
• the genes set limits but its largely the environment that determines where within those limits the organism lies
• e.g.
• temperature
• rainfall
• soil conditions
• pH
• food availability 
• Variation and selection
• variation and selection is necessary within  population
• selection can be natural or artificial.

Genetic Diagrams

Conventions

• Start by showing parental phenotypes then genotypes then gametes.
• A gene is represented by a single letter, upper case for dominant and lower cas for recessive
• Where a gene has more than two alleles, the gene has an upper case letter and the alleles are denoted in supercript.
• Where this is codominance the same convention applies

MONOHYBRID CROSSES

P1

Trait: Seed shaoe

Alleles: R- Round | r - Wrinkles

Cross: Round seeds x Wrinkles seeds

                  RR       x       rr

They would all be Rr

Genotype: Rr

Phenotype: Round

Genotypic ratio: all alike

phenotypc ratio: all alike

• homozygous dominant x homozygous recessive
• offring all heterozygous (hybrids)
• offspring called F1 generation
• genotypic and phenotypic ratio is all alike

F1 monohybrid cross

Traits: Seed shape

Allels: R- Round | r - Wrinkled

Cross:     Rr       x       Rr ( round seeds x round seeds)

Result:

RR                                   

Rr

Rr

rr

Gentotpes: RR, Rr, rr

Phenotype: Round and wrinkled

G ratio: 1:2:1 (RR:Rr:rr)

P.ratio: 3:1 (3 are rounded and 1 is wrinkles)

SEX LINKAGE

• a characteristic is sex linked if the gene that codes for it is found on one of the sex (X and Y) chromosomes

Convention

• when dealing with sex-linked characteristiccs in humans, the genotype of the male is XY (heterogametic) the female XX (homogametic)
• the sex linked allele is then denoted in supercript

Haemophilia A

• several factors are needed for blood to clot
• one of these is factor VIII - found on the X chromosome
• in it s recessive form the allele produces a non functional protein, blood takes longer to clot
• male only have on X chromosome
• if this has the recessive allele they will have haemophilia
• only having on allele for a characteristics is aka hemizygous

X^H= allele for normal factor VIII

X^h = allele for non functioning factor VIII

Cross between a carrier female and normal male
Results:

X^HX^H - normal female

X^Hy - normal male

X^HX^h - carrier female

X^hy - male with haemophilia

Duchenne muscular dystrophy (DMD)

• gene for muscle protein, dystrophin is on th X chromosome
• it is needed in muscular contration
• boys with te disease develop muscle weakness in early childhood and are usually wheelchair bound by age 10
• death often occurs due to complications of muscle degeneration (skeletal and heart uscle) by the early 20s

CODOMINANCE 

• this means thatthe homozygous dominant (AA) and the heterozygous dominant (Aa) both have the same phenotype = look the same
• however, in many cases when considering a pair of characters
• two alleles of a gene have an effect when they are present together
• when choosing symbols to represent co-dominant alleles it would be misleading to use a capital and lower case letters as this would suggest that the one with the cap was dominant
• instead, a capital letter is chosen to represent the gene and then two different superscript to represent the co dominant alleles
• a good example on incomplete dominance is found in snapdragons with respect to flower colour 

SICKLE CELL ANAEMIA

• the beta-strands of haemoglobin differ by 1 amino acid at position 6 (glutamic acid instead of valine)
• abnormal haemoglobin is de oxygenated and insoluble
• this deforms RBC making them inflexible and often sickle shaped
• they are unable to squeeze through capillaries
• after many oxygenated and de oxygenation cycles the RBC can be irreversibly damaged and sometimes destroyed
• when lodged in capillaries they can block the blood flow
• eventually organs esp heart lungs and kidneys become damage 

Sickle cell heterozygotes

• these individuals produce RBC with half of their haemoglobin normal and half sickled
• normal haemoglobin prevents sickling
• thus heterozygotes are symptomless carriers and at whole organism level this condition is considered to be recessive
• however, a molecular and cellular level because both alleles contribute to the phenotypes as observed in RBC it is co dominant

• Normal genotype: H^A H^A
• Sickle-cell gentoype: H^S H^S
• Symptomless heterozygous: H^A H^S

DIHYBRID INHERITANCE

• inheritance of two pairs of contrasting characters is known as dihybrid inheritance
• in pea plants:
• round seeds are dominant to wrinkled seeds
• yellow seeds are dominant to green seeds
• Y = allele for yellow seed coat
• y = allele for green seed coat
• R = allele for round seed shape
• r = allele for wrinkled seed shape
• when pure breeding plants with round, yellow seeds are crossed with plants that have wrinkled green seeds, all of the F1 generation had round yellow seeds
• These plants were allowed to self fertilise or were cross with each other and the resulting seeds were examined
• The expected ratio of 3 dominant to 1 recessive was found for both seed colour and for seed shape
• also, the inheritance of seed colour is independent of the shape of the seed and that nw combo of the characters appear in the F2 generations as well as the parental combination of the charcters

• Dihybrid inheritance is concerned with the inhereitences of the two genes at different loci
• genes that are on difference chromosomes (unlinked) are separated independently, and all combinations of two pairs of genes are possible
• a ratio of 9:3:3:1 is common when two factors are controlled y two independently sorted genes

EPISTASIS

• there are cases where different genes on different loci interact to affect one phenotype
• where one gene masks or suppresses the expression of another gene it is called epistasis
• they may either
• work antagonistically resulting in masking
• the homozygous presence of a recessive allele e.g. aa. may prevent the expression of another allele at a second locus.
• alleles at the st locus are epistatic to the alleles on the 2nd locus these are hypostatic
• work together in a complementary fashion
• recessive epistasis
• this can be seen in the inheritance of flower colour in salvia where two gene loci A/a and / on different chromosomes are involved
• a ration of 9:3:4 is observed
• this ration indicated recessive epsistasis
• the homozygous aa is epistatic to both alleles of gene B/ neither the allel or combiniion for purple  BB/b or the allele combination for pink b can be expressed i there s no dominant allele A present

Apoptosis

Events of apoptosis

• enzymes breaks down the cell cytoskeleton
• cytoplasm becomes dense with organelles tightly packed
• the cell surface membrane changes and small bits called blebs form.
• chromatin condenses and nuclear envelope breaks. DNA breaks into fragments
• the cells breaks into vesicles that are taken up by phagocytosis. the cellular debris is disposed of and does not damage any other cells or tissues

How it's controlled

• Apoptosis is controlled by cell signalling
• some from the inside the cell and some from outside
• these include cytokines made by cells of the immune system, hormones, growth factors and nitric oxide
• nitric oxide works by making the inner mitochondrial membrane more permeable to hydrogen ions
• it would prevent the formation of the proton gradient needed for ATP synthesis
• proteins are released into the cytosol
• these proteins bind to apoptosis inhibitor proteins and allow the process to take place

Apoptosis and development

• apoptosis is an important part of the tissue development. Extensive division and proliferation is followed by pruning through apoptosis. 
• Excess cells undergo apoptosis and their components are reused. Different tissues use different signals to induce it.
• Examples:the removal of ineffective or harmful T lymphocytes during the development of the immune system

How often does apoptosis take place?

• In children between 8-14 years old, 20-30 billion cells per day undergo apoptosis.
• in a year this is equivalent to the total body ass
• in adults the figure is around 50-70 million cells per day
• the rate of cell death should = the rate of mitosis
• however, not enough apoptosis leads to the formation of tumours. 
• too much leads to cell loss and degradation
• cell signalling helps maintain the right balance

Mutations - in depth

A mutation is change in the amount of or arrangement of the genetic material in the cell. They can be chromosomal mutations which involves changes to parts or whole chromosomes. Or, they can be DNA mutations which are changes to the nucleotide base sequences.

What causes mutations?

• During the process of replication, DNA is normally copied exactly so that the genetic material remains the same from generation to generation. However, occasionally changes can occur so that an organism may inherit altered DNA.
• The sudden change that can occur may produce an individual which is obviously different from the rest of the population. e.g. bacteria resistant to penicillin.
• Altered DNA molecules replicates the changed sequences so that the mutant genes are passed on to successive generations. - Such inherited changes are known as mutations.
• Mutation is another source of genetic variability n a population and it can occur spontaneously. 
• Mutant genes tend to be recessive but are transmitted in the normal way
• most mutations are harmful to the organism but some can be useful
• spontaneous mutations are essential for providing a new variation necessary for survival in a changing environment; in other words, it proves raw materials for evolutionary change
• The frequency with which one allele mutates to another is known as the mutation rate or mutation frequency and is measured by their occurrence in a population 
• mutations tend to be rare but the mutation rate can be speeded up by certain factors called mutagens or mutagentic agents which include x-rays, gamma rays and UV light which can damage DNA in various ways
• mutagens also include certain chemical substances which alter DNA by adding or deleting one or more bases in a sequences
• chemical mutagens include nitrous acid and 5-bromouracil

DNA mutations

• during protein synthesis, the sequence of ases in DNA is transcribed into a complementary sequence of bases in RNA which is then translated into a sequence of amino acids
• each sequence of three bases is a code for a specific amino acid, so that if the code is altered, this may result in an incorrect sequence of amino acids
• these changes would therefore result in non-functional proteins or proteins with a different function

Types of DNA mutations

• Point mutations - one base replaces another - also called substitutions
• Insertion/deletion mutations - one or more nucleotide pairs are inserted/deleted from a  length of DNA which causes frameshift
• A gene mutation in which a nucleotide is left out is called a deletion and a mutation in which a nucleotide is gained is called an insertion. 

• The effect on the sequence of amino acids from this small change in the DNA base is great.
• Every amino acid after the change ight be replaced by a different amino acid.
• both base deletion and base insertion have resulted in a shift so that all subsequent codons are altered.
• it is not surprising that mutation can result in the synthesis of a totally different protein

SICKLE CELL ANAEMIA

• The kind of point mutation that causes sickle cell anaemia is called substitution
• this is because the mutation is due to the substitution of one base for another just one place in the DNA molecule (thymine is replaced by adenine)
• the result of this is that the beta-chains of haemoglobin contains the amino acid valine at position 6 instead of glutamic acid
• Effect
• the haemoglobin is less soluble than normal haemoflobin and causes the RBC to become distorted into a sickle shape.
• this has many effects including severe anaemia and a clumping of cells which may lead to heart failure
• sickle-cell anaemia is often fatal before middle age

CYSTIC FIBROSIS

• 70% of the cases if cystic fibrosis the mutation is the deletion of a triplet base pair
• this deletes one amino acid out of a chain of 1480 in the normal polypeptide
• cystic fibrosis produces extra thick and very sticky mucus
• this can accumulate in the lungs and pathogens would normally be removed in the mucus by cilia  but the cilia are unable to move the mucus and thus causing infection - physiotherapy is required to get rid of it

ONCOGENES

• growth- promoting genes called protooncogenes
• these can be changed to oncogenes by point mutation so they cannot be turned off
• oncogenes promote unregulated cell division so this can lead to tumour. 

HUNTINGTON DISEASE

• Huntington disease results from an expanded triple nucleotide repeat
• the normal gene has a repeating CAG sequence above a certain threshold the protein alters sufficiently to cause the disease
• the symptoms are see later in life and include dementia and loss of motor control

Mutations w/ a neutral effect

• A change in the base sequence may produce no change in the organism if
• the mutation is a non-coding region of the DNA
• is a silent mutation - the change in the base does not alter the amino acid sequence
• the change in the organism is not seen as an advantage or disadvantage e.g. taste

Harmful/Beneficial effects:

• Early humans in Africa almost certainly had dark skin
• the pigment melanin protected them from the UV light
• they could however, still synthesise vit D from the action of intense sunlight on their skin
• humans with mutations resulting in paler skin would have been burned or suffered skin caner
• as humans migrated to more temperate areas sunlight was not intense enough to produce sufficient vit D by those with dark skins
• those with the pale mutations would have had an advantage (as a lack of vit D leads t rickets, narrow pelvis etc.)
• the Inuit peoples have not lost all their skin pigments although they dont live in an area with intense sunlight
• however, they eat a lot of vit D rich fish and seal meat.