Year 2- Chapter 11-manipulating Genomes

Question 1

What is a genome

Answer 1

This is the minimum quantity of genetic material that contains one copy of all the genes of an individual or of a population or a species.

Question 2

What is genomics

Answer 2

The application of the techniques of genetics and molecular biology to the mapping of genes on chromosomes and the sequencing of genes or complete genomes of organisms and viruses.

Question 3

How is genomes measured in different species and the units

Answer 3

Genomes of many species have been sequenced, which means that the whole of the base sequence is known. All of them have double-stranded DNA so their size is measured in base pairs (bp), kilobase pairs (kbp), or megabase pairs (Mbp).

Question 4

Which genomes are expressed well

Answer 4

Viral genomes are simple and prokaryotic genomes are smaller than eukaryotic genomes.
The control of gene expression in prokaryotes is much simpler than the control of eukaryotic genes. The genomes of eukaryotes have large quantities of DNA that are not transcribed or translated into proteins, but act to control gene expression. Also, eukaryotic genes have introns that are transcribed, but not translated.

Question 5

What are introns

Answer 5

A segment of DNA or RNA molecule which does not code for proteins and interrupts the sequence of genes

Question 6

What is gene expression

Answer 6

This is the process by which information from a gene is used in the synthesis of a functional gene product. These products are often proteins, but in non-protein coding genes such as tRNA or snRNA genes, the product is a functional RNA.
So there are genes that code for RNA molecules that conduct protein synthesis

Question 7

Structure of HIV

Answer 7

A single-stranded molecule of RNA with 5000 bases and nine genes

Question 8

E.coli structure

Answer 8

A single, circular molecule of double-stranded DNA with nearly 5 million base pairs and just over 4000 genes

Question 9

How is nuclear DNA organised

Answer 9

Nuclear DNA is eukaryotic organisms is divided into regions that have different functions. For example, there are structural genes that code for the assembly of amino acids to make polypeptides.

Question 10

How is a single gene in terms of its sequence of bases

Answer 10

Exons which are coding sequences (for proteins) are separated from non-coding introns. The gene for a beta polypeptide of haemoglobin has three exons and two introns and is 1605bp in length.

Question 11

What happens to the structural genes

Answer 11

These are transcribed as mRNA which is then modified and then translated.

Question 12

What happens to the other DNA structures

Answer 12

Other non-structural genes code for tRNA and rRNA. Some regulatory genes code for proteins that act as transcription factors and many code for forms of RNA that also control transcription and hence gene expression.

Question 13

What are promoters

Answer 13

These are control sequences found at the site of binding of RNA polymerase at the start of transcription. There are long lengths of DNA that separate structural and regulatory genes. These used to be considered as junk DNA but now they are considered to have a function.

Question 14

What are mtDNA and ctDNA

Answer 14

DNA in mitochondria are called mtDNA. DNA is chloroplasts are called ctDNA. These two organelles originated from prokaryotes and still possess what are essentially prokaryotic genomes with little non-coding DNA

Question 15

What are polymerase chain reactions

Answer 15

There are three steps to this:
Denaturation
Annealing
Extension

Question 16

What is denaturation

Answer 16

This occurs when hydrogen bonds between two polynucleotide strands are broken, thus separating the strands. This allows primers to gain access to the sequence of bases that are now exposed.
This process requires a temperature of 94 degrees centigrade for about 3 minutes.

Question 17

What is annealing

Answer 17

The temperature in the cycle now decreases to 50-65 degrees centigrade. At this conditions, hydrogen bonds can form between the two types of oligonucleotide primer and the complimentary DNA sequence of bases.

Question 18

What are the primers for?

Answer 18

The two primers are designed to bind to the region of DNA that is to be copied. Through complementary base pairing, one primer attaches towards the 5’ end of one strand and a different primer attaches towards the 5’ end of the other strand. These strands are antiparallel- one primer attaches at the ‘left’ of one strand and another primer attaches at the ‘right’ of one strand. The sequence of bases in each of the two primers is chosen carefully so that they bind to sequences just outside the region of DNA that is to be amplified (extended).

Question 19

What are primers important

Answer 19

Necessary to identify sites where synthesis will take place and because DNA polymerase functions by adding nucleotides to an existing piece of double-stranded DNA.

Question 20

What happens at the extension stage

Answer 20

DNA polymerase will add nucleotides to the primer. Therefore there will be an elongation as DNA polymerase builds up newly synthesised polynucleotide complementary to the template strands (similar to semi conservative replication). The DNA polymerase adds nucleotides to the 3’ end of the primer and synthesises the polynucleotide in the 5’ to 3’ direction. To do this dNTPs (deoxynucleotide triphosphates). This process requires a temperature of 72 degrees centigrade for 90 seconds.

Question 21

How are phosphodiester bonds between nucleotides formed in the extension stage

Answer 21

The hydrolysis of a bond between the first and second phosphate groups of each dNTP provides the energy for the formation of a phosphodiester bond between the newly added nucleotide and the growing strand.

Question 22

When and why are primers tagged with fluorescent molecules

Answer 22

So, DNA can be visualised and the progress of PCR monitored. This also allows the DNA to be analysed.

Question 23

What is Taq polymerase

Answer 23

The bacteria, Thermus aquaticus is the source of heat-stable DNA polymerase Taq polymerase. This is used in the extension stage of PCR.

Question 24

How many cycles can there be in the extension stage

Answer 24

Multiple
This is done by heating the new synthesised DNA molecules again. This separates the strands, making them available for copying again. Once, again this whole PCR cycle is conducted.

Question 25

What can a multiplex PCR do

Answer 25

It involves simultaneous amplifications of numerous DNA sequences in a single reaction mixture by using more than one pair of primers.

Question 26

Which compounds inhibit PCR reactions

Answer 26

Substances, associated with the stages of extracting and purifying the DNA hinder PCR reactions.
This includes ionic detergents, gel loading dyes and the enzyme proteinase K, used in extracting DNA from cellular material which, if left in the mixture, will break down polymerases. Similarly, certain substances present in blood can inhibit PCR, such as haemoglobin and the anti-clotting agent heparin.

Question 27

What is gel electrophoresis

Answer 27

This is a technique for separating and identifying substances in a similar way to paper and thin-layer chromatography.
Is used to analyse proteins and DNA and is carried out often on a paper or more often with gels.
This involves placing a mixture of molecules into wells cut into a gel, adding a suitable buffer solution and applying an electric field.

Question 28

What factors determine the number of charged molecules in the field

Answer 28

Composition of the gel:polyacrylamide gel are needed for proteins and agarose for DNA
Net charge of the molecule: negatively charged molecules move towards the anode (+) and positively charged molecules move towards the cathode (-); molecules with a higher charge move faster than those with less overall charge.
Size: smaller protein molecules and fragments of DNA move through the ‘holes’ in the gel faster than larger ones.

Question 29

How does electrophoresis of proteins work

Answer 29

The charge on proteins is dependent on the ionisation of the R groups on the amino acids residues. Some amino acids have R groups that are positively charged (-NH3+) whereas some of the amino acids are negatively charged (-COO-) or not charged at all.

Question 30

How does pH affect the electrophoresis of proteins

Answer 30

Whether these R groups are charged or not depends on the pH of the solution. When proteins are separated by electrophoresis the procedure is carried out at a constant pH by using a buffer solution. Usually the proteins are denatured in a reducing agent (mercaptoethanol) that breaks disulphide bonds. The proteins are often added to sodium dodecyl sulfate (SDS) which converts these proteins to negatively charged rod-shapes that move through the gel. The smaller proteins move faster through the gel and therefore travel further than larger proteins towards the anode.

Question 31

What else can gel electrophoresis be used for

Answer 31

It can be used to separate the polypeptides produced by different genes; also used to separate the variant forms of enzymes produced by different alleles of the same gene

Question 32

How can sickle cell anaemia (SCA) be tested in someone

Answer 32

The beta globin in haemoglobin molecules normally has the amino acid called glutamic acid- this has a non-polar R group so it is uncharged. In SCA, glutamic acid is replaced by valine acid, which has a R group that is charged. The two variants of beta globin can be separated by gel electrophoresis due to their difference in net charge. Haemoglobin in SCA, have a lower negative charge than normal haemoglobin. Due to this, they do not move as far through the gel compared to normal haemoglobin.

Question 33

What is the charge of DNA

Answer 33

All DNA fragments carry a small charge due to the negatively charged phosphate groups in the sugar-phosphate backbone of each polynucleotide.

Question 34

How does DNA electrophoresis work

Answer 34

The DNA fragments move through the gel towards the anode. The distance travelled by a length of DNA depends on its size- the shorter fragments travelled further.
The distance that the DNA fragments travel through the gel are determined by visualising the DNA with stains such as ethidium bromide and Azure A. The tracking dye moves through the gel slightly in front of the smallest DNA fragments so that the progess of electrophoresis can be followed and stopped before reaching the end of the gel.

Question 35

How to carry out an electrophoresis of DNA

Answer 35

1. Melt agarose gel in buffer solution
2. Insert a toothed comb at one end of the tank to make the wells to take DNA samples
3. Pour in the agarose gel.
4. Let the gel to set. Place electrodes at either end of the tank.
5. When the gel is set, pour in buffer solution and remove the comb.
6. Add blue dye to each DNA sample
7. Add DNA and dye mixture to the wells
8. Connect electrodes to the power supply.
9. When the blue dye in within 10 mm of the end of the gel, disconnect the power supply.
10. Pour away the buffer and add DNA stain (Azure A) for 4 minutes.
11. Rinse with water and analyse the fragments of the DNA, which will appear blue.

Question 36

How does capillary flow electrophoresis work

Answer 36

In this, the fragments of DNA (marked with fluorescent markers), are detected by a laser and a sensor that detects the fluorescence as each band passes towards the anode.

Question 37

What is principle of DNA sequencing

Answer 37

To find the order of the nucleotide bases along regions of DNA.

Question 38

Sanger sequencing or chain-termination method

Answer 38

Similar to PCR, when a newly synthesised DNA polynucleotide is being formed, free floating nucleotides are being arranged and bonded opposite to their complimentary bases. We do not know what base sequence of the polypeptide chain. So, in sanger sequencing, along with simple free-floating nucleotides, there are also nucleotides which have been marked with a fluorescent marker. These nucleotides terminate the synthesis of the DNA polynucleotide as soon as they are bonded to the rest of the nucleotide chain. Since, they have no particular order in which they bond with the polynucleotide fragment, these nucleotides stop the synthesis of the fragments at any point (in other words, they bond to the rest of the nucleotide chain randomly; at any point). This causes the fragments to be different sized. Different sized fragments are formed for every single of the four bases- so there are fluorescent terminating adenine nucleotides, thymine nucleotides, guanine nucleotides and cytosine nucleotides. Each base has been marked a different colour. The fragments can only contain one fluorescent nucleotide. The fragments are divided based on the type of terminating base they contain and poured into their base-labelled wells in the electrophoresis. During the process, the smaller fragments are able to move further and faster towards the anode, thus separating from the longer fragments. So, the fragments are ordered based on size. Each fragment will give off a colour which will indicate the base. So, while going through the size order, we can work out what bases each colour represents; thus the base sequence.

Question 39

What is shotgun sequencing

Answer 39

Each individual fragment of the DNA is read separately. Then, the overlaps are identified between the fragments. After this, the whole DNA sequence is put together.

Question 40

What have replaced Sanger sequencing

Answer 40

Next-generation sequencing.
These enable thousands of DNA molecules to be sequenced at the same time without the need for electrophoresis.
They are much faster than the traditional methods such as shotgun method and sanger method. Thus, the expenses in this process are reduced.

Question 41

What is nanopore sequencing

Answer 41

When DNA flows through the upper protein, a current is produced- since DNA is formed by ions; when there is a flow of ions a current is created.
Each base (A,T,C and G) alters current flow in a different way
Lower protein forms a pore through lipid bilayer
The change in current as each base passes through the pore is detected so the base sequence is determined.

Question 42

What is whole-genome sequencing

Answer 42

Comparisons of whole genomes allow scientists to find the location of the same or similar genes in different species and also provide data to analyse evolutionary relationships between species.

Question 43

What is Mycoplasma

Answer 43

By understanding the genome, researchers have been able to ‘knock out’ the genes in a simple organism to find out the minimum number required to keep the organism alive.

Question 44

What has gene sequencing allowed scientists to do

Answer 44

This has given to opportunities to design and make new molecules by writing completely new base sequences and inserting them into bacterial or eukaryotic cell using the techniques of genetic engineering. This is possible because they can use the genetic code to predict the amino acid sequence encoded by any length of DNA. This also enables t predict what shape the tertiary structure of a protein have.

Question 45

What advancements has genetical engineering enabled in medicine

Answer 45

Possible to involve enzymes in medicinal drug production
Changing the sequence of bases in existing genes to improve the way in which certain proteins work; such as various forms of insulins are available to treat diabetes
Redesign cellular systems so that organisms such as the genetically modified for the production of proteins, can work more efficiently.

Question 46

Definition of synthetic biology

Answer 46

This is the term coined for all the various applications of molecular and cellular biology; much of which relies on a knowledge of the structure and function of nucleic acids.

Question 47

What is epidemiology

Answer 47

This is the study of the spread of diseases in populations.

Question 48

How does a gene look like

Answer 48

When a nucleotide sequence is repeated in a DNA molecule, it is often considered a gene; with the variable number of repeats being considered as alleles.

Question 49

What is high-throughput sequencing

Answer 49

Sequencing machines containing 96 sets of capillary flow electrophoresis apparatus. These machines increased the speed at which different lengths of DNA synthesised by chain termination could be sequenced.

Question 50

What led to genetic profiling

Answer 50

The discovery of sequences that are repeated in DNA in structural genes and in regions between them led to methods of genetic profiling.

Question 51

What are minisatellites

Answer 51

Among first markers to be used were minisatellites, which are sequences repeated at various points along chromosomes. They can cut by restriction enzymes and the lengths of the fragments determined by gel electrophoresis.

Question 52

What are minisatellites replaced by

Answer 52

STRs, short tandem repeats, otherwise known as microsatellites. These are repeated sequences of nucleotides which are much shorter than minisatellites. They are regions of the genome composed of 2-5 nucleotides repeated 10-30 times. The number of repeats in any one STR is variable. This variability is caused by the inaccuracy of DNA polymerase in copying these regions during replication. The error rate in replication that leads to mutation is very low, but one effect is an increase in the number of these repeated base pairs. STR markers can be simple (identical length repeated sequences); compound (two or more adjacent repeated sequences) or complex (multiple length repeated sequences).

Question 53

DNA profiling in forensics

Answer 53

The STRs show a high degree of polymorphism, making them of particular use to the forensic scientist, and the STR regions are in non-coding DNA so there is no selective pressure against any of the alleles, the result that there is much variation between different people.
They tend to be four-five nucleotide repeats, as they are robust, suffer only low levels of environmental degradation and provide a high degree of error-free data. They represent discrete alleles that are distinguishable from one another, they show a great power of discrimination, only a small amount of sample is required due to short length of STRs.

Question 54

How can DNA be recognised through forensics

Answer 54

PCR is used to make copies of the DNA. Then, the sequences are separated by gel electrophoresis. The STRs are labelled with fluorescent dyes and detected by a laser scanner.
More modern analysis dispenses the electrophoresis stage and detects the fluorescence during PCR with four colours (e.g red, green, blue and yellow). A print out will form displqying the peaks.

Question 55

What is bioinformatics

Answer 55

Combining biological data with computer technology and statistics.
It builds up databases and allows links to be made between them. The database holds gene sequences, sequences of complete genomes, amino acid sequences of proteins and details of protein structures.

Databases can also hold data on macromolecules: their structures and functions, gene expression patterns, metabolic pathways and control cascades.
A future problem is collecting and organising data on the variety of proteins that are synthesised by eukaryotic cells.

Question 56

Examples of databases

Answer 56

The Genome OnLine Database (GOLD) is a comprehensive online resource to catalogue and monitor genetic studies worldwide. It provides up-to-date status on complete and ongoing sequencing projects along with a broad array of curated metadata.

Nucleotide sequences are held by the Nucleotide Sequence Collaboration between GenBank (USA), the European Nucleotide Archive and the Center for information Biology and DNA Data Bank in Japan. These databases compare sequences between the different organisations on a daily basis.

The database Ensemble holds data on the genomes of eukaryotic organisms. Among others it holds the human genome and the genomes of zebrafish and mice that are model organisms used a great deal in research.

Question 57

What can be done with the databases

Answer 57

Research and analysis
Retrieval(search) tool, BLAST, is an algorithm for comparing primary biological sequence information, such as the primary sequences of different proteins or the nucleotide sequences of genes.
Use to find similarities between sequences that they are studying
Sequences can be matched degrees of similarity can be calculated after a whole genome is sequenced. Very close similarity indicate close ancestry.
Organisms can provide useful models for investigating the way in which genes have their effect.
Provide vital information for vaccines.
In short, bioinformatics allows the comparison of genomes in different organisms to investigate evolutionary relationships. At one level, sequence data confirms the division of life into three domains.
At another level it shows the similarities between genes of organisms with very different phenotypes and the ways of life.

Question 58

What is genetic engineering

Answer 58

This term is often used to refer to the process of genetic modification by means that are not possible using selective breeding. It involves the removal of a gene or genes from one organism and placing them into another. At one extreme this involves transferring genes between species. But this can also involve taking a gene from an individual of a species and transferring it into other individuals of the same species.

Question 59

How is genetic engineering carried out

Answer 59

The DNA corresponding to a gene is obtained in one of a number of ways and inserted into a vector, such as a virus, plasmid, bacterial artificial chromosome (BAC) or liposome, which is used to transfer the gene into host cells. Alternatively, the DNA can be inserted directly into cells without using a vector.

The genetic code is universal so it is possible to make transfers between widely different species; for example, from a human into a bacterium.
All cells give this genetic code the same meaning so the protein originally encoded by the gene will be the same protein that is produced in any cell to which it is transferred.

Question 60

What are the reasons for wanting to transfer genes between genomes

Answer 60

Modifying bacteria and eukaryotic cells to make special chemicals that are only produced in small quantities by other methods
Making crop plants resistant to diseases, pests and herbicides
Making livestock resistant to diseases and pests
Improving the yields from crop plants and livestock
Improving the nutritional qualities of crop plants
Modifying animals to make human proteins for medicines that are difficult to obtain by other methods.
Modifying bacteria so they absorb and metabolise toxic pollutants.

Question 61

What is reverse transcriptase

Answer 61

An enzyme that catalyses the reverse of transcription. The enzyme uses RNA as a template to synthesise a polynucleotide using deoxynucleotide triphosphate molecules (dNTPs). The complementary DNA strand that is produced has a base sequence that is complementary t the sequence of bases on RNA. When a molecule of mRNA isolated from the cytoplasm of cells is used as the template,the cDNA has the same base sequence as the coding strand of the nuclear DNA from which it was formed in transcription. The advantage of this is that the cDNA has no introns, only exons.

Question 62

What is restriction enzymes

Answer 62

Are used to cut genes from lengths of DNA. They cut across both strands of DNA at specific sites known as restriction sites (as they are restricted to cut only at these sites).

Question 63

Where do restriction enzymes come from

Answer 63

From bacteria that are too dangerous or difficult to culture so the genes that code for them are removed and inserted into safer strains of bacteria that are modified to produce high quantities of them.

Question 64

What is the sequence of restriction enzymes

Answer 64

Each restriction site has a specific nucleotide sequence which is palindromic, so it reads the same in the 5’ to 3’ direction as in the 3’ to 5’ direction. They are present in bacteria to defend against attack by viruses; this is called restricting an infection by viruses.

Question 65

What are sticky ends

Answer 65

They cut DNA to give it free, unpaired ‘ends’. They will form base pairs with complementary sequences of bases. This is how a gene cut by restriction enzyme be inserted into a plasmid or into a viral DNA. Others cut straight across DNA to give blunt ends.

Question 66

What happens to a sequence of DNA once it is removed

Answer 66

Often necessary to produce large quantities of it. This is done by inserting the gene into bacteria and using the bacteria to replicate the gene.
Done by inserting the gene into a vector, such as plasmid or a virus, which then takes it into the bacterium.
The plasmid or viral DNA must have the same restriction site as the gene so that the same restriction enzyme can cut it to give a short length of complementary bases.
DNA cut by restriction enzyme can have a length of nucleotides added to give it sticky ends. Copies of the gene are mixed with plasmid.
Some plasmids will take up the gene by complimentary base pairing of sticky ends; other plasmids will simply reform without taking up the gene.
Hydrogen bonding between sticky ends attaches the two and the enzyme ligase is added to form the covalent phosphodiester bonds of the sugar-phosphate backbone of DNA.

Question 67

What are recombinant DNA (rDNA)

Answer 67

These are produced once the gene is inserted into a vector. The host bacteria are treated with calcium ions, then cooled and given an electric shock to increase the chances of plasmids passing through the cell surface membrane. This process is called electroporation.
The bacteria are now described as transformed because they contain foreign DNA.

Question 68

Can the recombinant DNA be formed every time? Are there any problems?

Answer 68

Some plasmids take up the foreign gene and some bacteria do not take up plasmids. There are variety of ways to identify the transformed bacteria from those that do not have the recombinant plasmids. Similar techniques are used to identify eukaryotic organisms that have been genetically modified.

Question 69

To identify the non-recombinant organisms, some techniques are used. Vectors may therefore contain marker genes such as:

Answer 69

Antibiotic-resistance genes: the foreign genes are inserted into these resistance genes so that they cannot be expressed; transformed cells are therefore sensitive to the antibiotic and untransformed cells are not the gene for green fluorescent protein (CFP), which is a small protein that emits bright green fluorescence in blue or UV light

The gene for beta glucuronidase (GUS), an enzyme from E.coli that converts colourless substrates into coloured products; it is used in plants to indicate that they have been genetically engineered.

Question 70

What does DNA polymerase do in bacteria

Answer 70

It copies the plasmids; the bacteria then divide by binary fission so that each daughter cell has several copies of the plasmid. The bacteria transcribe and may translate the foreign gene. If the bacteria produce a foreign protein they are described as transgenic. For transcription to occur in the host organism a promotor must be inserted with the foreign gene.

Question 71

What is a gene construct

Answer 71

The length of DNA that is constructed from promoter, foreign gene and marker gene, and maybe other genes as well, is known as a gene construct or just construct

Question 72

Why is bacteria useful in genetic engineering

Answer 72

Bacteria can make many copies of genes very quickly. This gives many copies that can be used in research.
Alternatively, the bacteria are grown in large quantities to make specific proteins, such as enzymes for the food industry.

Question 73

Why might genetic engineers not always prefer bacteria cells

Answer 73

It is not possible for the bacteria to carry out the complex post-translation processes of eukaryotic cells to cut, fold and modify proteins by adding sugars. As a result, genetic engineers might use yeast, plant or animal cells as the host cells for the production of proteins.

Question 74

What is the advantage of using yeast, bacteria and mammalian cells to produce proteins

Answer 74

Have simple nutritional requirements
Large volume of product are produced
The production facilities do not require much space
Processes can be carried out almost anywhere in the world

Question 75

What are GM crop plants

Answer 75

The first GM crop plants had genes to improve cultivation by incorporating pest and herbicide resistance. Pest resistance reduce losses to insect pests. Herbicide resistance allows farmers to spray herbicides during the growth of the crop to kill weeds that compete with the crop.
A recent development involves crops that are designed to improve human health.
Ex: GM rice known as Golden rice

Question 76

Soy bean plant example

Answer 76

Soy bean plants are highly susceptible to grazing by a variety of insect pests.
Losses to pests are estimated to run into billions of dollars each year. In response, a biotechnology company inserted a gene for pest resistance into its herbicide-variety known as Roundup Ready. The plants are modified to express a gene from the bacterium Bacillus thuringiensis which codes for a toxin (Bt toxin) that is poisonous to insect pests. The toxin binds to receptors located on the microvilli of epithelial cells in the gut of the larvae. It inserts into the membrane, causing formation of pores or ion channels and a water potential imbalance that eventually kills the insects.

Question 77

How to patent and transfer technology

Answer 77

Many biotechnology companies have patented the genetic modifications that they have developed. To gain an economic return for the years of investment in research and development, they charge farmers a higher price for GM seed than for non-GM seed. Farmers who grow GM crops such as Roundup Ready must buy the herbicide Roundup from the same company.
Increasingly new GM varieties will become available that combine that feature with others, such as pest resistance, high yield and efficient use of water.

Question 78

What are the objections to the development, use and release of GM organisms

Answer 78

Antibiotic resistance genes used to identify GM organisms could ‘escape’ and be transferred to pathogenic organisms, making it impossible to treat infections using such antibiotics
Herbicide-resistance genes could be transferred in pollen to weed species and lead to development of ‘superweeds’ that are resistant to herbicides.
An increase in infection by plant pathogens has been correlated with an increase in the use of glyphosate herbicides on herbicide-resistant crops; this has been particularly the case with alfalfa in the USA.
Foreign genes could be transferred to wild relatives of crop plants, or to non-GM and organic crops, so changing their genomes; this may ‘pollute’ those species.
Foreign genes could mutate once they have escaped and have unforeseen consequences; their insertion into the genome could activate or silence genes
Making crops resistant to herbicides enables farmers to use more herbicides than on non-GM crops; this increases costs to farmers
Farmers cannot keep seed for sowing for the following crop as GM crops do not ‘breed true’; this favours large-scale commercial farmers and not many farmers in developing countries
Genetic material from viruses used in genetic engineering could become incorporated with natural viruses so that an animal virus can infect humans or other animals or become more harmful; entirely new pathogenic viruses may evolve by this means
We are dependent on seven types of grain-producing crop plant, each of which is becoming genetically uniform and losing its genetic diversity; this means that the major food sources of the planet are increasingly dependent on our manipulation of their genomes to meet future environmental challenges.

Question 79

Is GM crops allowed everywhere

Answer 79

The growth of GM crops needs to be approved by authorities in the EU.
About half the countries, including France and Germany, have banned the growth of GM crops.
There is only one GM crop that has been approved for growth in the EU that is commercially grown. This is a type of maize that is resistant to the European corn borer, Ostrinia nubilalis.

Question 80

What about GM livestock

Answer 80

GM animals are divided into two groups. The first group has been genetically modified to enhance overall performance and the whole animal will become available for the food market. No GM animals have been approved by regulatory bodies for human food.

The second group of GM animals are transformed to produce specific substances in milk, egg or blood- so these can be used to treat diseases or serve as a medical research models.

Question 81

Proteins produced by transgenic animals

Answer 81

Sheep and goats have been genetically modified to produce human proteins in their milk

Human antithrombin is produced by goats: this protein is used to stop blood clotting

Human alpha-antitrypsin is produced by sheep: this is used to treat people with emphysema.

Question 82

What is ‘pharming’

Answer 82

The use of livestock to produce pharmaceuticals

The animals involved are known as ‘biopharm’ animals.

Question 83

Medicines produced by GM microorganisms

Answer 83

Insulin production for the treatment of diabetes
Human growth hormone to encourage growth in children with a deficiency of this hormone
Thyroid-stimulating hormone for treatment of thyroid cancer
Factors 8 and 9 for treating people who have a deficiency of one or other of these blood-clotting proteins
Vaccines
Monoclonal antibodies

Question 84

What proteins are genetically engineered

Answer 84

Restriction enzymes and ligases

Question 85

What are the advantages of using GM crops

Answer 85

Large scale production
Cheaper prices of the substances concerned
Production does not rely on other factors such as the availability of insulin from dead animals.

Question 86

What are GM pathogens- Mycobacterium tuberculosis

Answer 86

This has been modified to investigate its metabolism, drug resistance and the ways in which it causes disease. It has been modified so that it will grow more rapidly, making it easier to study in vitro. Such research also gives clues to ways in which vaccines can be developed and drugs produced for treatment.

Question 87

Why are viruses genetically modified

Answer 87

Have been genetically modified to deliver genes in gene therapy. Adenovirus is a very suitable vector as it will infect human and other mammalian cells; it is not species- or cell-type-specific.
These viruses have had two genes removed so that they do not replicate once they infect host cells. This removal gives space in the viral genome to insert a gene or genes.

Question 88

Adenovirus- when does it show up

Answer 88

The recombinant human adenovirus type 5 expresses enhanced green fluorescent protein (GFP) under the control of a promoter.
GFP is a protein originally isolated from Aequorea victoria, a bioluminescent jellyfish that fluoresces green when exposed to blue or UV light. Enhanced GFP is a GFP mutant with improved fluorescence and stability which is used as a marker. Therefore, a green glow in UV light indicates that the virus has successfully delivered its genes.

Question 89

Why is the Tobacco mosaic virus modified

Answer 89

To deliver the gene for a decapeptide hormone known as TMOF into the cells of crop plants.
The GM viruses are sprayed on crop plants and invade cells. The host cells transcribe and translate the gene to produce TMOF that inhibits the production of the enzyme trypsin by insect pests without having serious effects on the host plant.
The DNA sequence for TMOF was combined with that for TMV coat protein so that the decapeptide and coat protein are translated as one molecule. Trypsin cuts the peptide bond that joins to the two together so releasing active TMOF which then inhibits further production of trypsin by insects. After harvest,bthe leaves of the GM plants can be processed into a powder to be used as a spray to protect against insects such as mosquitoes.

Question 90

What are some ethical and practical problems in the GM pathogens

Answer 90

Proteins do not have to be extracted from animal sources or by collecting blood from many donors. The disadvantage of using bacteria to produce human proteins is that bacteria do not modify their proteins in the same way. It is much better, therefore, to use eukaryotic cells to produce human proteins.

Question 91

What are the potential hazards from releasing genetically modified organisms into the environment

Answer 91

Transgenic microorganisms do not compete well in the natural environment as they are engineered to produce substances that give them no advantage and they require much energy that is not readily available to produce these substances.
Containment facilities, such as filters on air conditioning and air locks on doors are fitted to prevent the escape of organisms
Lethal genes are added to the microorganisms so that they die if removed from the conditions of the culture.

Question 92

What is the CRISPR-Cas9 system

Answer 92

This occurs when DNA is cut in specific places, where the genome can be edited. One application of this system is to alter specific genes without changing other parts of an animal’s genome.
So, researchers can improve crops and livestock by replicating small genetic variations found naturally in different varieties of the same species.

Question 93

What is gene therapy

Answer 93

This is an application of the principle of genetic engineering. It involves several different methods to correct a genetic ‘error’.
Replacing a mutated gene that causes disease with a functioning version of the gene
Introducing a new gene into the body to help fight a disease
Repairing mutated genes by using an enzyme to edit the DNA and inserting a functioning gene.
Inactivating or ‘knocking out’, a mutated gene that is not functioning properly.

Question 94

What are the two types of cells that can be treated by gene therapy

Answer 94

Somatic cell gene therapy: somatic cells are body cells; all the cells in the body except gamete-forming cells and gametes. These cells all die when the individual dies so any genetic changes are not passed on to the next generation. However, gene therapy can be a treatment and maybe even a cure, for genetic conditions. Long-term treatments are possible by inserting a functioning allele into stem cells, such as those in bone marrow. It is most successful in cells that have a long life, such as those in the retina. It is less successful in cells that are replaced every few days, such as those in the airways and in the epithelium lining the gut.

Germ-line gene therapy: the germ line is the term given to cells that differentiate to form gametes. In mammals, the germ line forms fairly early in development and this group of cells populates the gonads: the ovaries and testes. These cells increase in number by mitosis and then some will divide by meiosis to form gametes. If a correctly functioning allele will divide into a fertilised egg, it means that all the cells formed by mitosis from that cell, once fertilised, will be genetically altered. The change is permanent and will be inherited by future generations.