Application of Reproduction and Genetics

eduqas alevel

what are stem cells?

undifferentiated cells that retain the ability to divide by mitosis an unlimited number of times to produce more stem cells or specialised daughter cells

what is differentiation?

the process where unspecialised stem cell becomes a specialised cell

What are totipotent stem cells?

can differentiate into any type of cells including embryonic and extra-embryonic (placental)

What are Pluripotent stem cells

Can differentiate into most types but not extra-embryonic

What are multipotent stem cells?

can differentiate into a limited range of cell types within a specific tissue or organ

what are embryonic stem cells?

unspecialised stem cells that can divide indefinately into any type of cell in the body

they can be taken from aborted fetuses and fertilised eggs from IVF

Why are embryonic stem cells useful?

-can be cloned for any cell

-can be used to study disease

-can study early stages of development- helpful to avoid miscarrage or treating birth defects and infertility

Problem with embryonic stem cells?

There arent enough, scientists want to use animal egg cells to generate embryonic stem cells and combine it with human cells. This has caused an uproar

advantages of stem cells

-speed of production

-large scale production

-production of genetically identical cells

-transplants to treat cancer

-test news drugs

disadvantages of stem cells?

-in mammals the technique is expensive and unreliable

-in plants disease, entry of pathogens may cause problems

-inadvertent selection of disadvantageous alleles.

What are Adult Stem cells?

They can only change into some stem cells in the body. This is because they can only specialise into certain types of cells as they have already started specialising

plant stem cells?

-found in meristems (roots and shoots)

-the vascular cambium is a layer of meristematic cells that give rise to xylem and phloem tissues

-able to differentiate throughout their entire life

How are stem cell used in tissue engineering

-Bone Marrow Transplants> treating blood related disorder

-Neurodegenerative disease> Parkinsons disease and Alzheimers by replacing lost neurons

-Other uses> treating type 1 diabete, macular degeneration, and spinal cord injuries

Ethical issues:

-controversial as it involves destruciton of a potential human life

-transported cells could become cancerous

Alternatives to ethical issues:

Iduced Pluripotent Stem Cells- adult cells reprogrammed to behave like embryonic stem cells- called inducing

What is DNA sequencing?

the process used to determine the precise sequence of nucleotides in a length of DNA

Who found DNA sequencing?

Fred Sanger> sanger sequencing method

How did the Sanger Sequencing method work

making a complementary template of a DNA and differing in length by one base pair. These are then separated according to length and, using gel electrophoresis the end base is identified and fluorescently dyed. Which allows the base sequence of DNA strand to be worked out

What was the human genome project and its age?

Finding the sequence of the entire human genome, finding the entire base sequence of genomes of simple eukaryotes, identifying new genes and their location, function and regulation, discuss ethical, social and legal issues

When did the human genome project begin?

1990-2000

How can the human genome project help medically

• Personalised drugs based on genomics

• Disease genes identified

• Pre-disposition to diseases identified so lifestyle advice can be given.

• Genetic disorders identified.

Ethical issues of Human genome project

-should parents have the right to know and not disclose pre-disposition to disease

-is electronically stored data saved

-do patients want to know about future issues

-is screening of embryos to choose the “best” ethical??

What sequencing method was used for the 100,000 genome project?

Next Generation sequencing which is much quicker

How does gel electrophoresis work

1- restriction enzymes cut DNA into smaller segments which are placed into wells in a porous gel and floats in a buffer solution between two electrodes

2-an electrical current is sent and the dna moves towards the postivive cathode

3- smaller DNA moves faster than larger DNA and so you can compare where DNA is the same in different sections of genes

Mosquitoes (anopheles gambiae)

sequencing the genome of this malaria vector allows scientists to study the causes insecticide resistance which can be used to develop new chemicals and strategies to reduce mosquito populations and prevent disease transmission

Malarial parasite (plasmodium)

researchers sequence plasmodium to identify targets for new drugs and vaccines

Primates (chimpanzees)

genomic comparisons between humnas and primates help determine evolutionary relationships and common ancestry.

model organisms

genomes of organisms like E. coli, yeast and roundworms are sequences to provide reference points for basic biologival processes and medical research

ethical issues of genetically modifying mosquitoes

-may alter ecology of area

-could lead to extinction of mosquito by man

Steps of genetic finger printing

1. Collection - A biological sample such as blood, saliva, hair roots, or skin cells is obtained

2. Extraction - DNA is extracted from the biological sample

3. Amplification - PCR is used to amplify (make many copies) of the DNA fragments. Amplifies the regions of introns with different numbers of STRs. This is useful as the sample may be very small.

4. Digestion - Restriction enzymes cut the DNA (digest) at specific recognition sites leaving the VNTR sequences intact.

5. Separation (Gel Electrophoresis): DNA fragments are placed in a gel and an electric current is applied. DNA is negatively charged and moves toward the positive electrode; smaller fragments move faster and further than larger ones, separating them by size.
   Southern Blotting: The DNA is transferred from the fragile gel to a more durable nylon or nitrocellulose membrane. The resulting pattern is visualized using techniques like X-ray film or UV light.
   Hybridization: DNA probes (short, single-stranded sequences labelled with a radioactive or fluorescent marker) are added to bind to complementary VNTR sequences.
   Visualization: The pattern of bands is revealed using X-ray film (for radioactive probes) or UV light (for fluorescent probes)

How does PCR work?

1- Uses enzymes to seperate the double strand of DNA with high temperatures (break H bonds)

2- Primers anneal to sperated strands as temperatures are reduced due to complimentary base hybridisation from the primer, determining which section gets copied

3- DNA polymerise catalyses the synthesis of complementary dna strans

why is genetic fingerprinting unique

the specific length and number of VNRTs are highly variable

What is genetic fingerprinting used for?

-matching DNA

-paternity/ maternity

-genetic relationships

-measuring variability within genetics

-selective breeding

What are the main used of genetic engineering

-production of useful substances

-production of plants or animals with more useful characteristics

-gene therapy

What is Recombinant DNA

DNA that contains genes that do not normally exist together

What is a plasmid

small loop of doubled DNA often found in bacterial cells, carries extra genes and is often used as a vector to transfer new genes

What are restrcition enzymes

cuts DNA at a specific site, which is determined by base sequence

What are Sticky ends

when DNA is cut they produce single stranded over hangs.

The sequence allows other fragments to anneal to complementary sticky ends. Hydrogen bonds form between them

DNA Ligase

catalyse the formation of phosphdiester bonds between the sugar-phosphate backbone of each DNA strand

Transformation of Bacteria

the uptake of DNA into bacterial cells. This usually involves the transfer of a DNA Plasmid as a Vector that carries the new of altered gene.

What can bring about transformation

-calcium chloride treatment

-electric shock

-heat shock

What is reverse transcriptase

an enzyme that synthesises DNA from RNA. Can use mRNA to reproduce a cDNA strand and the mRNA can be isolated, which is easier then islolating the whole genome

Marker genes

An additional gene inserted into a plasmid that is used to aid in the identification of host cells that have taken up the desired gene.

How is recombinant DNA made

1- gene and vector are isolated

2- splicing occurs> gene is inserted into vector

3- transformation of vector into cells

4-growth

5- selection of cell/ organism with inserted gene

What problems does using cDNA solve?

-locating the gene

-restriction enzymes cutting the gene into non functional fragments

-introns

How to use mRNA as a starting material

1- extract mature mRNA (only introns) from desired cell.

2- use reverse transcriptase to producse a single stranded cDNA

3- use DNA polymerase to produce double stranded DNA

4- Inster double stranded DNA into vector

Concerns over genetic engineering of bacteria

-possible transfer of antibioctic resistance genes to pathogenic bacteria

-possible transfer/ activation of oncogenes by using fragments of human DNA

Ti plasmid

-a common plant cloning vector found in soil bacteria

-this bacteria infects plants and a section of the Ti plasmid becomes incorporated into the host cell genome, producing a gall or tumorous lump

-this can be used gto genetically modify plants

GM plant example : soya

GM soya has been produced that is resistant to herbisides

advantages to GM crops

-solution to food shortages

-improved nutrients

-longer shelf life

-higher yields

-pesticide resistance

concerns with GM crops

-potential transfer of resistant genes to wild populations via pollen

-reduction of biodiversity

-unknown effect of eating new protein in crop

-produce resistant insects

What is gene therapy and what are the two methods

treatiung genetic disease by replacing defective genes

-somatic cell therapy (no inhertiance )

-germ line therapy (inhertiance)

Example of gene therapy: Duchenne Muscular Dystrophy

-a recessive sex linked form of Muscular Dystrophy affecting 1 in 3500 live male births

-it is caused by a mutation in the dystrophon gene resulting in failure to produce dystrophin which is an important structure componnent of muscle tissue> wasting of muscles, wheelchair bound by teens and life expectancy of 27

-a drug called drisaperson is used as a “molecular patch” to bind and mask the mutated exon, meaning it is ignored in protein synthesis. Called exon skipping

challenges and limitations of gene therapy

-viral vectors can cause inflammation and serious side effects

-effects aren’t permanent

-efficient and safe delivery to correct cells is difficult

-ethical concerns about altering human gene pool

Ethical issues with the Human Genome project

-provides personal information about people

-who owns the genes and other pieces of DNA?

-parents must be fully informed before fetal screening

-should parents have the right to let their children be tested for adult diseases

-concerns that people will choose specific characteristics for their babies