1. Applications of Genetics
Learning Objectives:
Human genetics
Pedigree Analysis
Genetic Screening
Genetic Counseling
Gene Therapy
Human Genome Project

2. 2. Plant and animal breeding
Artificial Selection
Cloning
3. Recombinant DNA Technology and its applications
4. DNA fingerprinting and its applications
5. Implications of genetic manipulation: the potential benefits, hazards and ethical issues.

3. Human Genetics
Pedigree Analysis
Pedigrees = Family Trees
One of the central tasks of the human geneticist
Pedigree analysis is the construction of family trees
Family history information is often collected at major family gatherings
A pedigree is used to trace inheritance of a trait over several generations.

4. Symbols used in pedigree charts:

5. Inheritance of Haemophilic trait in the Royal Family

6. Three primary patterns of inheritance:
autosomal recessive
autosomal dominant
sex-linked (X-chromosomal)

7. Autosomal Recessive Pedigree
Recessive: neither parent has the characteristic phenotype (disease) displayed by the child
Autosomal: Gene is on one of the autosomes (Chromosomes 1-22).
Phenotype only occurs under homozygous condition

8. Autosomal Dominant Pedigree
Dominant: Affected individuals can appear in every generation
Autosomal: Gene is on one of the autosomes (Chromosomes 1-22).
Phenotype occurs under both homozygous and heterozygous conditions

9. Sex-Linked Pedigree
X-linked: The trait is preferentially seen in males, who are homozygous. Females are heterozygous "carriers"
Most X-linked traits are recessive.
Inheritance of red-green color blindness, an X-linked, recessive trait:

11. Color blind male is father of "carrier" daughters and normal sons.
Carrier daughters have 50% chance to have color blind sons.
Color blind male x carrier female can produce color blind daughters
Can you determine a pedigree for Red-Green Colorblindness?

12. For more information, please refer to the following website:
Pedigree analysis problem set available at

13. Genetic screening
Diagnosis of inherited diseases before symptoms occur by finding abnormalities in the genes or chromosomes.
Some examples:
sickle cell anaemia
cystic fibrosis
phenylketonuria (PKU)
Down’s Syndrome
Most of these tests performed on cells removed from the fetus (pre-natal diagnosis) and even from a pre-implantation embryo.

14. Ultrasound
One of the simplest and easiest genetics screening tests
Evaluate the physical characteristics of the fetus which may help predict certain abnormalities
Measurements of various body parts can be used to suggest an increased risk of Down Syndrome
look for congenital defects such leaky heart valves, and birth defects such as cleft lip or club foot.
Ultrasound is also used to guide the physician during amniocentesis and chorionic villus sampling (CVS).

15. An example of ultrasound image

16. Amniocentesis
Remove fetal cells from amniotic fluid 14 to 22 weeks after pregnancy.
Culture the cells and look for
chromosome abnormalities (e.g., the three number 21 chromosomes of Down syndrome);
certain enzymatic defects (e.g., an inability to metabolize galactose, hence milk);
the sex of the fetus.

17. Risks:
To the mother: slight discomfort, adverse reaction to medications used, vaginal bleeding or cramping, and infection.
To the foetus: inadvertent puncture is small
To the pregnancy: chance to miscarriage very low (not more than 1/200)

18. Couples who may wish to consider amniocentesis include:
1. Women 35 years of age and over
2. Parents who have had a child with Down's syndrome or other chromosome abnormality.
3. Couples who are known carriers of a chromosome rearrangement
4. Couples who have had a child with a malformation of the brain or spinal cord

19. Chorionic villus sampling (CVS)
Procedure:
Suck out placental cells by a tube inserted through the abdomen or vagina.
Advantages:
No need to perform cell culture
Can be performed earlier in pregnancy (10-12 weeks)
If an abortion is to be performed, it is a simpler process early in pregnancy

20. Drawbacks:
More risky than amniocentesis
Increase the chance of miscarriage (approximately 0.8%)
Minor complications such as vaginal bleeding or cramping occur more frequently following CVS than amniocentesis
Infection
Transverse limb defects in infants can be resulted
Involve the absence of the distal structures of the limb
May result in the vascular system disruption of the limb
Risk for transverse limb defects following CVS is approximately 0.03%-0.10%

21. Couples who may wish to consider CVS include:
Women 35 years of age and older
Parents who have had a child with Down's syndrome or other chromosome abnormality
Couples who are known carriers of a chromosome rearrangement
Couples who have a family history of a genetic condition for which testing is available

22. Common genetic diseases diagnosed by CVS
Chromosomal abnormalities such as Down’s, Klinefelter’s and Turner’s syndromes can be identified by karyotype analysis.
The cells can be cultured for DNA analysis that helps diagnosis of
1. Cystic fibrosis
2. Huntington’s chorea
3. Thalassemia

24. Genetic Counseling
Genetic counselors are trained persons who help individual and family to
comprehend the medical facts, including the diagnosis, probable course of the disorder, and the available management;
appreciate the way heredity contributes to the disorder, and the risk of recurrence in specified relatives
understand the alternatives for dealing with the risk of occurrence
choose the course of action which seems to them appropriate in view of their risk, their family goals, and their ethical and religious standards, to act in accordance with that decision
make the best possible adjustment to the disorder in an affected family member and/or the risk of recurrence of that disorder.

25. Who Should be Referred?
Families in which one or more members have a serious birth defect or genetic disease.
Families who have a child with multiple congenital anomalies, serious developmental delay, or an unexplained abnormality of growth.
Families in which more than one close relative has the same disease, e.g., mental retardation, deafness, blindness, cancer, early heart attacks, or schizophrenia.
Couples who have had repeated stillbirths or a stillborn baby with birth defects.
Couples who are close blood relatives, e.g., first cousins.

26. Common issues discussed during genetic counseling
Making a diagnosis
Investigate the family history for previous cases of genetic diseases through pedigree analysis
calculate and explain the risk of having affected children
explain the cause of the disease
Quality of life
The quality and likely length of an affected child’s life will be discussed
Availability of treatment, support groups and financial help will also be discussed.
Effects on other family members will be discussed

27. Options:
These include contraception, sterilization, adoption, artificial insemination to avoid the husband’s genes being passed on or IVF using a donor egg if the problem lies with the woman.
Prenatal diagnosis is an option if the couple are willing to consider abortion of an affected fetus.
CVS, amniocentesis and abortion must be discussed.
The reliability of tests such as DNA analysis, ultra-sound scanning must be explained.
Possibility of IVF and gene therapy

28. Ethical issues in genetic counseling
During postnatal counsel
Risk evaluation, preventive options, gene therapy, testing of all family members, confidentiality within the couple
During prenatal counsel
Option of pregnancy termination, selective implantation, sex selection
Problems of Eugenics (The birth of designer baby)

29. Gene Therapy
A new method for treating genetic diseases by replacing faulty genes with normal genes or adding normal genes if they are absent.
Two types of gene therapy:
Somatic cell gene therapy - the insertion of genes into body cells which function only in the treated person and are not passed on to the offspring.
Germline gene therapy - the insertion of genes into cells that are involved in reproduction which can be passed on to the future offspring.

30. Diagrammatic Illustration of Gene Therapy

31. General Procedure of Gene Therapy:
Isolate and clone the normal gene.
Introduce it into the chosen human cells with the help of a safe and efficient vector.
The cells may have to be isolated from the body first, corrected and then replaced.
Easier for blood diseases such as sickle cell anaemia because the cells that make blood cells can easily be removed from bone marrow and replaced.
The final problem is to make sure that the gene is expressed normally.

32. Common Vectors Used:
Viruses (e.g. Retrovirus) carrying the required gene
Liposomes containing the cDNA clone
Microinjection and electroporation- Direct injection of donor DNA into the cell

33. Applications of Gene Therapy e.g. Cystic fibrosis
Cystic fibrosis affects the epithelial cells of the body, but the life-threatening problems mainly affect the lungs.
Lung and trachea epithelial cells are therefore the initial targets for gene therapy. The aim is to get the gene into the cells so that it can make the normal protein, known as CFTR
The cDNA clone is enclosed in a specially designed vector.
Adenovirus infecting the respiratory tract is currently used as the vector for the CFTR gene
The vector does not insert its DNA into the host DNA. If the cell divides, the new DNA is not replaced at the same time so it eventually becomes diluted.
The treatment may only be effective for a few weeks until the epithelial cells die, but it will be easy to repeat the treatment at regular intervals.

34. Advantages Disadvantages Advantages and Disadvantages of Gene Therapy
Addition of normal allele will alleviate symptoms
Only somatic cell therapy allowed
Avoid need for constant medication
Not a permanent cure as cells shed
Extend life expectancy
Viral vector may give inflammation problems
Possibility of DNA entering cells other than target cells with unknown effects

35. Ethical issues of gene therapy
Germline therapy is controversial because it opens up the whole field of eugenics.
The same technique can be used to add genes for desired characteristics. (For example, the American public has demonstrated a desire for enhanced growth of normal children with demands for human growth hormone)
Germline therapy is controversial because the change can be passed on to the children of the treated person and all subsequent generations.
Do we have the right to alter the genome of the future generation?

36. Human Genome Project (HGP)
The goals of the project are to:
identify all the approximately 30,000 genes in human DNA
determine the sequences of the 3 billion chemical bases that make up human DNA
store this information in databases
develop faster, more efficient sequencing technologies
develop tools for data analysis, and
address the ethical, legal, and social issues (ELSI) that may arise from the project.

37. Applications of HGP
Molecular Medicine
Improved diagnosis of disease
Earlier detection of genetic predispositions to disease
Rational drug design
Gene therapy and control systems for drugs
Pharmacogenomics "custom drugs"

38. 2. Microbial Genomics
New energy sources (biofuels)
Environmental monitoring to detect pollutants
Protection from biological and chemical warfare
Safe, efficient toxic waste cleanup
Understanding disease vulnerabilities and revealing drug targets

39. 3. Risk Assessment
Assess health damage and risks caused by radiation exposure, including low-dose exposures
Assess health damage and risks caused by exposure to mutagenic chemicals and cancer-causing toxins
Reduce the likelihood of heritable mutations

40. 4. Bioarchaeology, Anthropology, Evolution, and Human Migration
study evolution through germline mutations in lineages
study migration of different population groups based on female genetic inheritance
study mutations on the Y chromosome to trace lineage and migration of males
compare breakpoints in the evolution of mutations with ages of populations and historical events

41. 5. DNA Forensics (Identification)
Identify potential suspects whose DNA may match evidence left at crime scenes
Exonerate persons wrongly accused of crimes
Identify crime and catastrophe victims
Establish paternity and other family relationships
Identify endangered and protected species as an aid to wildlife officials (could be used for prosecuting poachers)
Detect bacteria and other organisms that may pollute air, water, soil, and food
Match organ donors with recipients in transplant programs
Determine pedigree for seed or livestock breeds
Authenticate consumables such as caviar and wine

42. 6. Agriculture, Livestock Breeding, and Bioprocessing
Disease-, insect-, and drought-resistant crops
Healthier, more productive, disease-resistant farm animals
More nutritious produce
Biopesticides
Edible vaccines incorporated into food products (Biopharming)
New environmental cleanup uses for plants like tobacco

43. Ethnical problems
Fairness in the use of genetic information by insurers, employers, courts, schools, adoption agencies, and the military, among others.
Who should have access to personal genetic information, and how will it be used?
Privacy and confidentiality of genetic information.
Who owns and controls genetic information?
Psychological impact and stigmatization due to an individual's genetic differences.
How does personal genetic information affect an individual and society's perceptions of that individual?

44. More information on HGP is available at
Dolan DNA learning center (2002), Gene Almanac, [Online] Available at
(The Human Genome WebQuest)

45. Selective Breeding of Animals and Plants
Aims:
To select desirable traits and remove undesirable traits of plants and animals through artificial selection
Two types of breeding involved in artificial selection:
Inbreeding
Outbreeding

46. Inbreeding – mating of closely related individuals
Advantages: Create pure line over time
Better adapted to steady environment.
For example
Parental genotypes: Ffgg x Ffgg
Gametes: Fg , fg Fg , fg
F1: FFgg Ffgg Ffgg ffgg
(pure breeding)

47. Disadvantages of inbreeding
Increasing the chance for recessive genes to be homozygous
Most recessive genes are undesirable traits
Leads to a high frequency of defects present at birth
Reduce the genetic variability vigour and fertility of a population.

48. Outbreeding - mating of individuals that are unrelated
Advantages:
Progeny (offspring) are heterozygous and the bad recessive genes are masked by normal dominant alleles.
Hybrid vigour – Progeny are tougher, more fertile and have a greater chance of survival
Produces variation/heterozygosity; on which natural selection can act.
Disadvantage:
No more pure line exists

49. Examples
The cross of larger, non-sweet-tasted tomato with small, sweet-tasted tomato.
The cross of high-resistant to pest and disease crop with higher-yield crop.
Parental genotypes: FFgghhIIjj x FFGGHHiiJJ
Gametes: FghIj FGHiJ
F1: FfGgHhIiJj
Higher chance of heterozygosity in F1
Result in hybrid vigour

50. The hybrids are stronger and bigger in size than their parents
male
parent
female

51. Artificial Insemination for Domestic Animals
Placing semen within the female reproductive organs through any means other than sexual intercourse.
Particularly useful with:
Conditions preventing vaginal ejaculation
Sperm with low motility (movement)
Cervical mucus that is hostile to sperm
Presence of sperm antibodies

52. Procedure:
Intracervical insemination (ICI) OR Intrauterine insemination (IUI)
ICI - Fresh ejaculate (semen) is placed directly into the cervix (opening to the uterus) by using a syringe and cannula (long, slender tube).
IUI - "Washed" semen is placed directly into the uterus via a catheter through the cervix.

53. Success Rates
ICI - 2% or higher chance of pregnancy per cycle
IUI - Higher probability of pregnancy than ICI. Generally, % chance of pregnancy per cycle Rates are higher when AI is accompanied by superovulation using fertility drugs.

54. Typical Procedure Protocol:
Pre-procedure:
Thorough evaluation of both male and female for suspected causes of infertility
Determination of drugs to be used (or not) and timing of treatment cycle
Monitoring by ultrasound and blood work to evaluate ovarian response to drugs and reduce potential side effects
If drugs result in ovarian hyperstimulation syndrome, treatment cycle will be discontinued immediately.
In case of no drug use, patient may be required to monitor own cycle with ovulation predictor kits and other methods induction of ovulation by HCG , if necessary

55. During procedure:
Male partner must produce a semen sample, usually in the clinic (may be collected at home in some cases), or donor semen must be available
Female partner is inseminated while lying down, feet in stirrups, on examination table
Female may be advised to refrain from strenuous physical activity for 24 to 48 hours after insemination

56. Advantages of using artificial insemination
Sperm from one male can be used for many females;
One ejaculate can be used for several inseminations;
Speeds up selective breeding;
Saves cost of keeping male and travelling for mating
Less stress than mating;
Allows genetic testing

57. Disadvantages of using artificial insemination
Danger of inseminating too many females with one male causing inbreeding;
With consequent genetic problem;
Undetected genetic weakness
Requires expertise;
Cost of vet.;
Sperm damage in storage;
Not successful in some species ;

58. Diagram showing how sperms are collected

59. Selective breeding of animals – sperm bank (for domestic animals and human)
Sperms are collected and frozen in liquid nitrogen for further use.
Endangered animal species, e.g. giant pandas can be bred
Sperm from one male (with good quality) can be used for many females
Infertility treatment in human

60. Selective breeding of animals –embryo transfer breeding (for domestic animals and human)
Procedure:
Injections of hormone to stimulate and ovulation in the animals that you want to get the embryos from.
The donor is inseminated at normal time
Seven days later, rinsing out the uterus to extract the embryos and ova (unfertilised, fertilised or degenerate)
Isolation of the good embryos using a microscope and then transfer into the recipient animals e.g. surrogate.

61. Applications:
Domestic animal e.g. cow
Produce up to ten or more progeny per year from their best cows.
Profit from the increased sale of quality genetics without losing the bloodlines.
Extend the productive life of some older cows, incapable of carrying another calf by producing further progeny through the use of embryo transfer.
Conserve the genetics in their herd through the uses of, embryo freezing for, export, domestic sale or future transfers on their own farm.
Infertility treatment in human