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Question 1

Advantages of Asexual reproduction

Answer 1

-Quicker.
-Optimal conditions shared between parents and offspring.
-Only need one parent.

Question 2

Disadvantages of asexual reproduction

Answer 2

-No genetic diversity.
-Does not allow for natural selection.
-Whole population is susceptible to change.
-Little variation in the population.

Question 3

Clones

Answer 3

-Genetically identical copies.
-Produced through asexual reproduction in which the nucleus is divided mitosis.
-Two identical copies of the DNA are created, and then separate into genetically identical nuclei.

Question 4

Vegetative propagation

Answer 4

-Asexual reproduction in plants.
-Eg plants produce:
-Runners (on surface),
-Rhizosomes (below ground)
-Suckers (stems that grow from the roots of a plant).
-Bulbs (An underground stem from which a series of leaf bases grow).
-Tubers (Another underground stem).
-Can occur artificially through cuttings, grafting or tissue cultures.

Question 5

Cuttings

Answer 5

-Take a healthy shoot.
-Remove all but a couple of leaves to reduce transpiration.
-Cut stem at a slant between nodes and dip in root powder with auxins.
-Place in soil or compost.
-Also can be done with plant roots, dormant twigs, and leaf cuttings.

Question 6

Micropropagation

Answer 6

-A group of cells (explant) are removed from the parent plant (often meristem tissue as free from viruses).
-Explant is sterilised and placed on a nutrient culture such as agar gel, containing suitable concs of glucose, phosphates and plant growth hormones.
-Cells divide into a group of totipotent stem cells (callus).
-Callus divided to produce small clumps of undifferentiated cells.
-Clumps stimulated to grow, divide, and differentiate into different plant tissues.
-The different tissues are determined by growth mediums containing different concentrations of auxin and cytokinin/
-Plantlets transported to a greenhouse to be grown.

Question 7

Advantages of plant cloning

Answer 7

-Desirable parent plant can be chosen.
-Plants mature at the same time.
-Can be carried out where sexual reproduction is not possible (eg bananas)
-Quick.
-New plants are free from viruses.
-New plants all uniform in phenotype, which makes them easier to grow and harvest.

Question 8

Disadvantages of plant cloning

Answer 8

-Labour intensive.
-Risk of contamination.
-Lack of genetic diversity.
-Expensive

Question 9

Tissue culture

Answer 9

-A series of techniques used to grow cells, tissues or organs from a small sample of cells or tissue.
-Carried out in a nutrient medium under sterile conditions.

Question 10

Reproductive cloning (animals)

Answer 10

-Produces large numbers of genetically identical animals.
-Useful for high-yield farm animals or genetically modified animals.
-Either embryo splitting or SCNT.

Question 11

Embryo splitting

Answer 11

-Mammals can produce two identical offspring if an embryo splits very early in development.
-IVF occurs between the egg of a high value female and the sperm of a high value male.
-A zygote is produced from IVF and allowed to divide via mitosis.
-Cells are separated and allowed to continue dividing.
-Each small mass of cells is placed into the uterus of a surrogate mother.

Question 12

Somatic Cell Nuclear Transfer (SCNT)

Answer 12

-The only way to clone an adult (first was Dolly the sheep in 1996).
-The nucleus is removed from an egg cell (enucleation).
-Normal body cell (somatic cell) from the adult to be cloned is isolated.
-Body cell is fused with the empty egg cell by applying an electric shock, which also triggers the egg cell to start developing.
-Cell undergoes mitosis, and the small ball of cells (embryo) is placed into the uterus of a surrogate mother.

Question 13

Therapeutic cloning

Answer 13

-Non-reproductive.
-New tissues and organs can be grown as replacements.
-Eg for skin grafts, repair of malfunctioning organs, and potential growth of new organs.

Question 14

Advantages of artificial animal cloning

Answer 14

-Can produce large amounts of animals with desirable characteristics.
-Retains desirable characteristics.
-Uses genetically identical embryos and tissues, meaning scientific research can investigate the effects of genes and hormones without the impact of genotypes.
-Testing medicinal drugs and cloned tissues avoids using animals for testing.
-Can produce cells or tissues for donors.

Question 15

Disadvantages of artificial animal cloning

Answer 15

-Lack of genetic variation.
-Animals may all be susceptible to one disease.
-Animals cloned for desirable characteristics may have a poorer quality of life.
Success rate of adult cell cloning is very poor, and is expensive.
-Ethical issues regarding the creation and destruction of life.

Question 16

DNA profiling procedure

Answer 16

-DNA is obtained from the individual.
-DNA is digested by restriction enzymes, which split it into fragments at specific recognition sites.
-Fragments are separated by gel electrophoresis.
-This creates a banding pattern.
-The DNA compared against the individual’s is treated with the same restriction enzymes and subjected to electrophoresis.
-The banding patterns of the DNA samples can be compared.

Question 17

Types of DNA analysed during DNA profiling

Answer 17

-Originally (1978) used fragment length polymorphism analysis.
-Today used short tandem repeat (STR) sequences of DNA, which are highly variable short repeating lengths of DNA.
-Each STR is polymorphic, but has a small number of alleles in the gene pool.
-Thirteen STRs are analysed each to avoid the chance of all STRs being present in both people.

Question 18

Applications of DNA profiling

Answer 18

-Forensics, through establishing DNA left at crime scenes to determine or refute innocence.
-Settlement of parental disputes, as half of the STR fragments will come from the mother and half from the father.
-Analysis of disease to determine proteins present.

Question 19

Early DNA research

Answer 19

-In the 1970s the structure of DNA, and the sequence of base triplets for amino acids, were known.
-Scientists worked from transcribed mRNA from genes rather than DNA.
-This process was extremely slow and only suitable for short genes.

Question 20

Sanger’s sequencing approach

Answer 20

-Uses a single strand of DNA as a template for four experiments in separate dishes, each containing a solution with the four bases, plus DNA polymerase.
-A base labelled with a radioactive isotope was added to each dish. These were modified so that, once they were incorporated into the synthesised complementary strand of DNA, no more bases could be added.
-Thousands of DNA fragments of varying lengths were produced (as each added base cut off the strand), and passed through a gel by electrophoresis.

Question 21

Sanger’s experiment results

Answer 21

-Smaller fragments travelled further through the gel during electrophoresis.
-The base at the end of each fragment was read by its radioactive label.
-Eg if the first one-base fragments has thymine at the end, then the first base in the sequence is T.
-If the two base fragments have cytosine at the end, then the sequence is TC.

Question 22

Dis/advantages of Sanger’s method

Answer 22

-Efficient.
-Safe.
-Used to sequence the genome of a phage virus.
-Bases had to be counted off one by one from the bands in the gel.
-So very time consuming and expensive.

Question 23

Agarose gel electrophoresis (setup)

Answer 23

-Used to separate different sized fragments of DNA for identification and analysis.
-Uses an agarose gel plate covered by a buffer solution in a tank. This is heated and then cools with a comb placed at one end.
-Once the gel is set the combs are removed to leave wells at one end of the gel.
-DNA samples are digested with restriction enzymes to cut them into fragments (at specific recognition sites), and then added using a pipette to the wells (with a loading dye).
-A well is also filled with DNA-size standard.
-Electrodes are set up at either side of the tank so an electric current (18V) can pass through the gel for 6-8 hours.

Question 24

DNA size standard

Answer 24

-Contains DNA strands of known lengths.
-Can give a reference for length of samples during electrophoresis.

Question 25

Movement of DNA samples in electrophoresis

Answer 25

-DNA is negatively charged due to its many phosphate groups. -Therefore moves towards anode (positively charged electrode). -Shorter strands move further due to having less mass. -The current is shown to be running by the presence of air bubbles from the electrodes.

Question 26

Ethidium bromide

Answer 26

-Dye stains DNA samples to show them under fluorescent light. -Binds to DNA, so can damage DNA in cells (mutagen).

Question 27

Separating proteins

Answer 27

-Similar process to electrophoresis, but involves a charged detergent such as sodium dodecyl sulphate (SDS) to equalise the surface charge of the molecules and allow the proteins to separate as they move through the gel according to molecular mass. -Can sometimes be separated according to mass and then by surface charge. -Can be used to analyse haemoglobin proteins to diagnose conditions such as sickle cell anaemia (patient has hg S rather than hg A).

Question 28

DNA probes

Answer 28

-A DNA probe is a short (50-80 nucleotides) single-stranded length of DNA that is complementary to a section of the DNA being investigated. -May be labelled using a radioactive (anneals to 32P in phosphate groups and is exposed with photographic film) or fluorescent (UV light) marker. -Used to locate a specific gene for genetic engineering. -Used to identify the same gene in genomes from different species for genome comparison. -Identify the presence or absence of a specific allele for a genetic disease.

Question 29

Microarrays

Answer 29

-A number of different DNA probes on a fixed surface. -Can reveal the presence of mutated alleles that match the probes as the sample DNA anneals to complementary base probes. -DNA must be broken into smaller fragments, and can be amplified using PCR. -Reference and test DNA samples are labelled with fluorescent markers. Where a test and reference both bind to a probe the scan reveals fluorescence of both colours.

Question 30

Annealing

Answer 30

-The binding of a probe by complementary base pairing to be the piece of DNA.

Question 31

Cloning DNA

Answer 31

-The gene to be sequenced is isolated using restriction enzymes. -DNA is inserted into a vector (bacterial plasmid) and then into an E Coli bacterium host. -E Coli divides many times, meaning the plasmid is copied many times.

Question 32

DNA sequencing machine

Answer 32

-First automated machine developed in 1986. -Fluorescent dyes instead of radioactivity were used to label the terminal bases. -Dyes glowed when scanned with a laser beam, and the light signature was identified by a computer.

Question 33

High throughput sequencing

Answer 33

-Faster, cheaper methods of sequencing. -Pyrosequencing involves the synthesis of a single strand of DNA complementary to the strand to be sequenced. -The bases produced are detected by light emission produced during their synthesis (by the enzyme luciferase) and can be sequenced by the light produced. -This research has led to the creation of bioinformatic, which enables the storage of the huge amounts of data required.

Question 34

Applications of gene sequencing

Answer 34

-Genome comparisons between species that allow for identification of characteristics genes code for (eg genes for insulin in humans and pigs, FOXP2 coding for speech). -Evolutionary relationships through similar genomes (high throughput techniques show 99% similarity between chimps and humans), can also detect most recent common ancestors. -Variation between individuals due to 'single nucleotide polymorphisms (SNPs). -Predicting the amino acid sequences of proteins due to knowing what gene codes for a protein.

Question 35

SNPs

Answer 35

-Around 0.1% of human DNA is not shared with other humans. -This means there are three million places on the DNA lengths where DNA sequences can differ due to random mutations. These are called SNPs. -Some can alter a protein, or the way a piece of RNA regulates the expression of another gene.

Question 36

Epigenetics

Answer 36

-Methylation is the addition of a methyl group (CH3) to a substrate. -Methylation of chemical groups in DNA plays a major role in regulating gene expression. -Methods to map this methylation in human genomes can help researchers understand the development of certain diseases and whether they may develop in genetically similar individuals.

Question 37

Synthetic biology

Answer 37

-Designs and builds useful biological devices and systems. -Includes biotechnology, molecular biology, and biophysics. -Goals include biological systems that store information, provision food, and improvement of human health.

Question 38

Polymerase chain reaction (PCR)

Answer 38

-Developed in 1983 to amplify the amount of DNA in a sample, enabling it to be analysed. -PCR is the artificial replication of DNA. -Differs from natural replication in that it only occurs in short sequences (10,000 base pairs), and requires primer molecules and heating and cooling to separate the strands.

Question 39

PCR ingredients

Answer 39

-DNA polymerase (duplicates DNA) from Taq from Thermophilus Aquaticus (suitable at high temperatures). -2x primers. -Free nucleotides. -Original DNA strand to be replicated. -Thermocycler, computer controlled machine that varies temperatures for set time periods.

Question 40

Applications of PCR

Answer 40

-Detecting mutations/oncogenes in a sample from a patient of a disease. -Identifying viral genome in DNA's host cells. -Forensic science, as small quantities can be amplified. -Research of extinct animals by amplifying DNA.

Question 41

Primers

Answer 41

-A piece of single stranded DNA complementary to the specific target sequence at the 3' end of each replicated DNA strand. -Used in polymerase chain reaction.

Question 42

Denaturation

Answer 42

-First stage in polymerase chain reaction. -The sample of DNA is mixed with DNA nucleotides, primers, magnesium ions and Taq DNA polymerase. -Reaction mixture is heated to 96C for thirty seconds. -Hydrogen bonds between base pairs break, and denature the DNA into two single strands.

Question 43

Annealing

Answer 43

-Second stage in the PCR. -Mixture is cooled down to around 68C. -The primers bind (anneal) to the 3' end of the single separated DNA strands. -This gives a small section of double-stranded DNA at the end of each single stranded molecule. -This means the copying process can begin.

Question 44

Amplification (Polymerisation)

Answer 44

-Final step of PCR. --The Taq DNA polymerase enzyme molecules now bind to the end with double-stranded DNA. This occurs at 72C, meaning the DNA remains single stranded. -Taq DNA polymerase catalyses the addition of DNA nucleotides to the single stranded DNA molecules, starting at the end with the primer and going in the 5 to 3 direction. -The the Taq DNA polymerase reaches the other end of the DNA molecule, a new double strand of DNA is generated. -PCR is repeated around thirty times to give around 1 billion copies of the original DNA. -This takes around 3 hours.

Question 45

Capillary electrophoresis

Answer 45

-Runs electrophoresis through a capillary rather than a gel. -Much faster.

Question 46

Genetic engineering/recombinant DNA technology

Answer 46

-Combining of DNA from different organisms (genetic modification). -Genes are isolated from one organism and inserted into another organism using vectors.

Question 47

Stages in genetic engineering

Answer 47

-Isolation of DNA containing the required gene. -Insertion of the DNA into a vector (eg plasmid). -Transformation, the transfer of DNA into a recipient cell. -Expression of the gene in the recipient.

Question 48

Isolation

Answer 48

-Methods of isolating the DNA containing the required gene include: -Obtaining mRNA from cells, then making cDNA copies using reverse transcriptase. -Locating the gene with a probe and cutting it out with restriction enzymes. -Using PCR to amplify the gene from genomic DNA. -Synthesising a copy using an automated polynucleotide synthesiser.

Question 49

Reverse transcriptase

Answer 49

-An enzyme made up from a group of retrovirsuses. -Used to turn viral RNA into cDNA so that it can be transcribed in the host cell into proteins.

Question 50

Using reverse transcriptase (insulin)

Answer 50

-RNA extracted from B cells in the Islets of Langerhans. -Extract mature mRNA coding for insulin. -Reverse transcriptase forms a single-stranded complementary copy of DNA (cDNA) using the mRNA template. -DNA polymerase (using primers) forms DNA from cDNA. -This forms a double stranded copy of human insulin, which can be inserted into plasmids using a ligase enzymes.

Question 51

Restriction enzymes

Answer 51

-In bacteria to protect from attack by phage viruses. -Cut up DNA of viruses. -Prokaryotic DNA is methylated at recognition sites to protect from restriction enzymes. -Used in molecular biology to recognise specific sequences in a length of DNA and cleave to molecule there.

Question 52

DNA Ligase

Answer 52

-An enzyme used to join DNA fragments. -Catalyses condensation reactions that join the sugar groups and phosphate groups of DNA. -Used in DNA replication, and PCR, and to insert sections of DNA into a plasmid.

Question 53

Insertion (1)

Answer 53

-Second stage in genetic engineering. -Plasmids (circular molecule of DNA) are obtained from organisms such as bacteria and mixed with restriction enzymes that will cut the plasmid at specific recognition sites. -The cut plasmid has exposed unpaired nucleotide bases, called sticky ends, or recognition sites.

Question 54

Insertion (2)

Answer 54

-Free nucleotide bases complementary to the sticky ends of the plasmid are added to the ends of the gene to be inserted. -The gene and cut plasmid anneal at the sticky ends. -This is catalysed by DNA ligase.

Question 55

Vector

Answer 55

-A vector (eg a circular piece of DNA called a plasmid) is used to transport DNA into the host cell. -Plasmids are useful because they nearly always contain antibiotic resistance genes. A large amount of DNA can be inserted into them. -Plasmids usually come from bacteria such as E. Coli. -Weakened (attenuated) viruses can also be used.

Question 56

Transformation

Answer 56

-Plasmids are reintroduced into the host cell of bacteria. -However, DNA does not easily cross the recipient cell's plasma membrane. -Bacteria are subjected to alternating hot and cold periods in the presence of calcium chloride, meaning their walls and membrane will become more porous. -This is because the positive calcium ions surround the negatively charged parts of the DNA molecules and the phospholipids in the cell membrane, reducing repulsion. -Can also occur through electroporation, in which a high voltage pulse is applied to the cell to disrupt the membrane. -Small pieces of gold or tungsten can be coated with DNA and shot into plant cells (gene gun). -Plasmids can be inserted into bacterium that can infect hosts and naturally insert its genome.

Question 57

Containment of bacteria

Answer 57

-Bacteria with recombinant genes in also have the gene that synthesises a nutrient removed. -This means that they cannot survive outside of the laboratory where they are given than gene.

Question 58

Potential hazards of gene manipulation

Answer 58

-Genetically modified crops may be toxic to animals. -Herbicide resistant genes thought to spread herbicide resistance into weeds. -Many object to the use of animals for medical testing, or the welfare of genetically modified animals.

Question 59

Gene therapy

Answer 59

-The insertion of a functional allele of a particular gene into cells that contain only non-functioning alleles of that gene. -Means the individual will produce a functioning gene and no longer have the symptoms of a genetic disorder. -Improved from knowledge gained by the Human Genome Project. -Attempts have been made to use interference RNA to block translation of harmful genes.

Question 60

Somatic gene cell therapy

Answer 60

-Metabolic orders (cystic fibrosis) occur when an individual inherits two recessive alleles of a particular gene. -The differentiated cells lack the protein product of that gene. -Functional alleles for the gene can be inserted into specific cells, so they produce the normal protein. -These alterations do not pass to the parent's offspring.

Question 61

Liposomes

Answer 61

-Alleles are packaged within spheres of lipid bilayer (liposomes). -These can be sprayed into the noses of patients, and will pass through the plasma membrane/nuclear envelope of cells in the respiratory tract. -Insert into the host genome, meaning the host cell will express the desired protein. -Cells lining the respiratory tract are replaced every two weeks, so treatment must be repeated.

Question 62

Viruses in gene therapy

Answer 62

-Viruses can be genetically modified to encase the functioning allele (instead of viral alleles), and infect the recipient cells with the allele. -However, viruses provoke an immune response, and may lead to the patience becoming immune to the virus. -In some tests patients developed Leukaemia, as the virus disrupts a gene involving regulating cell division.

Question 63

Germ line gene therapy

Answer 63

-Altering the genome of gametes or zygotes. -Means all cells of the individual will be altered, and their offspring may inherit the foreign alleles. -Ethically controversial, as it is changing the genetic makeup of many people who cannot give consent. -Insertion of genes may disrupt the expression of other genes.