Karnival Pendidikan Melaka 2009

Karnival Pendidikan Melaka 2009

Biology MoleculeToxicologyChemistry & Food TechnologyPlant Tissue CultureMicrobiology

Biology MoleculeToxicologyChemistry & Food TechnologyPlant Tissue CultureMicrobiology

Applications of Molecular Biology Techniques in Biotechnology

Industry

Amelia Kassim

Nurhuda Jalil

Nurul Arneida Husin

‘Education Fair 2009’ Melaka Institute of Biotechnology

Molecular biology is the study of biology at a molecular level.

INTRODUCTION

One of the most basic techniques of molecular biology to study protein function is expression cloning

Melaka Institute of Biotechnology

Plant Tissue Culture

Microbiology

Disciplines relationMarket intelligence

BioinformaticProteomic

What is DNA cloning?

When DNA is extracted from an organism, all its genes are obtained

In gene (DNA) cloning a particular gene is copied (cloned)

• To "clone a gene" is to make many copies of it• Act of making many identical copies of gene• Gene can be an exact copy of a natural gene • Gene can be an altered version of a natural gene

What is Gene cloning?

How is DNA cloned?

Cell-based DNA cloning Cell-free DNA cloning (PCR)

Clone in dividing cells

First cycle

Primer oligonucleotide

DNA replication

Denaturation, annealing

DNA sequence to be amplified

Second cycles

20 – 30 cycles

Amplified DNA sequences

Clone in PCR

Some Molecular Biology Techniques1. DNA / RNA extraction2. pDNA purification3. Polymerase chain reaction (PCR)4. Expression cloning5. Gel electrophoresis6. Gel documentation, and image analysis7. Southern blotting8. Northern blotting9. Western blotting and immunochemistry10. cDNA / genomic DNA library construction, and its screening 11. Southern, and Northern Hybridization techniques12. Auto-radiography 13. Primer designing 14. PCR-techniques 15. PCR product Cloning 16. Construction of transformation vectors 17. Biolistic PDS He/1000 mediated plant transformation 18. Agrobacterium mediated genetic transformation19. Analysis of the transgenic plants 20. Microscopy21. Cultivation of bacteria / Microbiology, 22. preparation of glycerol stocks23. Generation of ESTs24. Analysis of DNA, cDNA / ESTs sequences using online and offline bioinformatics tools 25. Promoter isolation and characterization ++++++++++++++++++++++++++++++

When you are working in

Molecular Biology Lab

Melaka Institute of Biotechnology

Gel-Documentation

RT-PCR

Spectrophotometer

Melaka Institute of Biotechnology

Gel Electrophoresis

Applications of Molecular Biology in Biotechnology Industry ?

Some Examples:

1. Genetic engineering of plants / Animals/ Microbes

2. DNA Fingerprinting

3. Novel (Blue rose)

4. Exploitation of natural resources plant product development (Genes for Perfumes in Plants)

5. Gene Cloning - Insulin and other drugs (HIV?) production?

6. Criminals monitoring based on DNA

7. Fruits with insulin, Milk with Antioxidants / vitamin A ++

Metabolic Engineering

Power of Metabolic Engineering

Genetically modified

DNA FINGERPRINTIN

G TECHNOLOGY

Prepared by:

Nurul Arneida Husin

Molecular Bioinformatics

DNA???• Deoxyribonucleic acids

• Building blocks of life.

• Nucleic acid that contains within it the genetic code that makes up the blueprint of the biological development of all plant, animal and some virus life forms.

• Our unique marker that can uniquely identify us.

• No two organism will ever have the same DNA, except in rare cases of twins.

Deoxyribonucleic acid (DNA) is a nucleic acid that contains the genetic instructions used in the development and functioning of all known

living organisms and some viruses.

By 1953,

Crick and Watson

had built a model which incorporated all known features of DNA, and

proposed the double helix structure which is

commonly referred to as the Watson-Crick model of

DNA.

Crick, Watson and Wilkins were awarded Nobel prize

for this work in 1962.

A-T

G-C

T-A

C-G

C-G

G-C

A-T

BASE PAIRING

A- ADENINE

T- THYMINE

G- GUANINE

C- CYTOSINE

DNA FINGERPRINTING?????

•Known as DNA PROFILING.

•Techniques for analyzing and comparing DNA from separate sources, to identify any organism (human, plant, animals, microbes etc) from their biological materials.

How to perform DNA fingerprinting?

• Study and Characterization of Plant Morphology• Selection and Identification of Plants• Isolation of Genomic DNA• Selection of Specific Primers• PCR (Polymerase Chain Reaction) Amplification• Gel Electrophoresis Analysis• DNA Analysis of Species-Specific Marker• Development of Plant Database

DNA Fingerprinting Process

Raw materials

Isolation of Genomic DNA

Loading PCR product

Selection of Primer/s

Adding PCR Component and tested samples

Run PCR

PCR product

Run Gel Electrophoresis

Unique DNA Fingerprint profiles

Data analysis

What is DNA Marker Technology?

DNA Marker Technology is a set of technologies such as Random Amplified Polymorphic DNA (RAPD), Amplified Fragment Length Polymorphism (AFLP), and Microsatellites.

Application to herbal industry?

• DNA Marker Technology has found application in fingerprinting genotypes, in determining seed purity, in systematic sampling of germplasm, and in phylogenetic analyses.

• DNA based techniques have been widely used for authentication of plant and herbal species of medicinal importance. This technology is very helpful for the identification of the variety within species or species within the generic group.

• Global trend leading to increased demands of medicinal plants for pharmaceuticals, phytochemicals, nutraceuticals, and cosmetics.

• Therefore, this service technology is applicable for those in herbal business sector to make quality assurance and correct identification of the starting materials to ensure reproducible quality of herbal medicine, which contributes to its safety and efficacy.

• It is because herbal medicinal products may vary in composition and properties, unlike conventional pharmaceutical products, which are usually prepared from synthetic, chemically pure materials by means of reproducible manufacturing techniques and procedures.

• This application of markers can be explored for standardization of botanicals formulation containing medicines.

Molecular Markers in Herbal Drug Technology

• Herbal drug technology….is used for converting botanical materials into medicines, where standardization & quality control with proper integration of modern scientific techniques & traditional knowledge is important.

• Authentication of plant species of medical importance.

• Genotyping & quick identification of botanical samples.

• Detection of adulteration/substitution• Marker assisted selection of desirable

chemotypes• Medicinal plant breeding• Applications in foods & nutraceuticals

Roles of Molecular Markers in Herbal Drug Technology?

Molecular Markers in Essential Oils Production

Roles of Molecular Markers in Increasing Essential Oil Production…..

• Breeding strategy• To identify right cultivar for increasing

essential oils production.

• DNA Marker study will provide information on ways of enhancing the economics of essential oil production.

Microbiology World and You

MICROBIOLOGY UNIT

What is Microbiology?

Micro - too small to be seen with the naked eye

Bio - life ology - study of

Why study Microbiology?

Microbes make the world go round. Microbiology lies at the heart of the biosciences. There is hardly an area of medicine unaffected by microbes and microbes are essential for the environmental cycles that regenerate our planet.

Fields in Microbiology

Medically related fields: Medical Microbiology Immunology Epidemiology Virology Molecular Pathogenesis

General and Applied Microbiology Environmental Microbiology Molecular Microbiology Industrial Microbiology   Biodiversity Bioremediation Food Microbiology

What Are My Options?

Microbiologists can specialize in a variety of areas. From the food to the space industries, and everywhere in between, there are many opportunities for microbiologists!

- Biochemist - Environmental Scientist- Biotechnologist - Bacteriologist - Virologist - Immunologist - Mycologist - Teacher

FUNGIVIRUS

What kind of work does an industrial microbiologist do? The application of scientific and engineering principles to the

processing of materials by microorganisms (such as bacteria, fungi, algae, protozoa and viruses) or plant and animal cells to create useful

products or processes.

Responsible for the discovery, development, or implementation of certain processes and the quality of resultant products:

- Antibiotics/Antimicrobials- Vaccines- Health care product - Food & beverages produced my microbiology activity

What kind of work does an medical microbiologist do? The study of microorganisms including bacteria, viruses, fungi and parasites

which are of medical importance and are capable of causing diseases in human beings.

Work with the diagnosis and management of infectious diseases in hospitals and in the community and in preventing people at risk.

What kind of work does an food microbiologist do? Food Industry - Oversee and optimize the conversion of raw materials into finished products - Ensure product quality and safety - Carry out new product development in a very competitive industry Public Service - Develop / enforce legislation and regulations related to food production and safety - Inspect and evaluate food production facilities - Improve post harvest technologies at home and abroad

Technical Sales and Marketing - Participate in sales and marketing of ingredients, equipment and packaging materials - Interface between suppliers and the food industry - Provide scientific and technical support

Where Does a Microbiologist Work and With Whom?

Microbiologists work as integral members of interdisciplinary teams in laboratories of hospitals, clinics, universities, health care, research organization, industry (food and drink, pharmaceutical, toiletry, and biotechnology companies) and government.

The following lists provide a brief sampling of the kind of jobs and work environments

Sample Career Titles Sample Work Settings Lab Technician Private Industry

Research Technician Hospitals

Clinical Microbiologist Private Industry

Microbiologist Education

Clinical Trials Manager Contract Research Organization

Lab Research Analyst Medical Research Laboratory

Plant Tissue Culture LABORATORY

MELAKA INSTITUTE OF BIOTECHNOLOGY

ACTIVITY

1. Research and Development of various types of medicinal, ornamental and forest plants species.

RESEARCH OFFICER

2. Mrs. Azurin Hanim bt. Jamaluddin.3. Mrs. Rabiatul Adawiyah bte Mohd.

Yusof.

INTRODUCTION

What is plant Tissue Culture???

Plant tissue culture is a practice used to propagate plants under sterile conditions, often to produce clones of a plant

INTRODUCTIONPlant tissue culture relies on the fact :

Any plant cells have the ability to regenerate a whole plant(totipotency).

Parts of plants can be used to generate a new plant are such as :

Single cells Protoplasts (plant cells without cell walls ) Pieces of leaves. Roots Stem

Explants was given with the required nutrients and plant hormones included on culture media in order to propagate.

OBJECTIVETo produce/propagate :

The exact copies of plants which have desirable traits.

Quickly mature plants.Multiples of plants in the absence of

seeds or necessary pollinators to produce seeds.

Whole plants from plant cells that have been genetically modified.

Plants with greatly reduced chances of transmitting diseases, pests, and pathogens.

Plants which have very low chances of germinating and growing, i.e.: orchids

METHODS IN TISSUE CULTURE1.Media Preparations 2.Explants Preparations3.Culturing the explants

4.Subculturing the plantlets

5.Transplanting (Acclimatization)

Shoot multiplication

Root induction

Shoot induction

Callus induction

MEDIA PREPARATIONS

Addition of Nutrients

Weighing Sucrose Measure pH

Addition of gelling agentsMedia cooking Autoclave

PREPARATIONS OF EXPLANTS (selection and sterilization)

Culturing the explants

Subculturing the plantlets

Transplanting (acclimatization)

Transfer to nursery (hardening)

Chilli

Sambung Nyawa

Orchids

Misai Kucing

Orchids

Chilli

Kacip FatimahRose

Our products

Misai Kucing

Sanserviera Trifasciata Teak

Nilam

Limau Kasturi

Sundal Malam

Chilli

Orchids

Misai Kucing

FOOD &

CHEMICAL TESTING

LABORATORY

MELAKA INSTITUTE OF BIOTECHNOLOGY

‘Bridging Research with Industry’

ACTIVITY1. Research and Development of Natural Plant and

Herb2. Food and Chemical Analysis

Research Officer3. Ms.Ong Pei Thing, AMIC4. Cik Sabrina Hj Md Kassim5. En. Musalman Rambeli

Natural Plants and Herbs

1. Screening of various local plants and herbs

for antioxidant properties.

2. Production of antioxidant-rich extract from plants and herbs.

3. Development of nutraceutical and cosmeceutical products from potential plants.

4. New formulation of Biopesticide on Vegetable Farming.

Offers various types of food testing:

i. Protein vii. Energy

ii. Total Fat viii. Benzoic Acid

iii. Moisture ix. Sorbic Acid

iv. Ash x. Saturated Fat

v. Carbohydrate xi. Cholestrol

vi. Vitamin A,C,D and E

Chemical Testing

i. Steroids

ii. Heavy Metals

iii. Minerals

iv. Essential Oil

Water Testing

i. Biochemical Oxygen Demand (BOD)

ii. Chemical Oxygen Demand (COD)

iii. pH

iv. Dissolve Oxygen (DO)

v. Total Soluble Solid (TSS)

vi. Total Dissolve Solid (TDS)

vii. Heavy Metals

viii. Minerals

Gas Chromatograph / Mass Spectrometer (GC/MS)

i. Used in detecting and analyzing the chemical compound typically from organic compound.

Used in analytical and biochemistry to separate, identify and quantify compounds.

a measurement of the dissolved sugar-to-water mass ratio of a liquid.

Efficient method to remove solvent from sample by evaporating usually used with plant and herbs methanolic extracts.

Toxicology Unit

Bridging research with industry

Melaka Institute of BiotechnologyLot 7, MITC City, Hang Tuah Jaya, 75450 Ayer Keroh, Melaka.06-231 3266

Biology Fair

Kolej Matrikulasi Melaka

Established in early 2007

With 2 research officers

Small laboratory with few basic equipments – Elisa plate reader for cytotoxicity testing, centrifuge, 160L fridge, laminar flow cabinet for mammalian cell sterile working area, fluorescence and inverted microscope and CO2 incubator for culturing cell.

In intention to expand the function of this lab, we have proposed a project to MOSTI to develop a mutagenicity testing facility in accordance to OECD GLP principle.

On December 2007, MIB has been granted this project which cost about RM4 million for 3 years.

This facility for mutagenicity testing with GLP compliance is estimate accomplished on 2010.

Research for toxicology appointed as one of collabrators with

local/international research institute; UiTM – toxicology profile for newly synthesis

stilbene

Services - for local/international companies manufacturing food products and supplements

short term studies – offer services for in vitro toxicology testing

provide toxicology profile testing

Mammalian cell line based;i. In Vitro Cytotoxicity and Test (MTT Assay) and mode

of cell death determinationii. In vitro Mammalian Chromosome Aberration Test

(OECD 473)iii. In Vitro Mammalian Cell Gene Mutation Test (OECD

476)iv. In Vitro Sister Chromatid Exchange Assay in

Mammalian Cells (OECD 479) v. Single Cell Gel Electrophoresis Assay (Comet Assay)

Bacteria cell based;i. Bacterial Reverse Mutation Test (Ames Test) (OECD

471)ii. Umu Test

Toxicology (from the Greek words toxicos and logos) is the study of the adverse effects of chemicals on living organisms.

It is an inter-disciplinary science that integrates the principles and methods of many fields: chemistry, biology, pharmacology, molecular biology, physiology and medicine. It study the symptoms, mechanisms, treatments and detection of poisoning, especially the poisoning of people.

Risk assessment - attempts to determine, probabilistically, outside the laboratory, the likelihood of an adverse effect based on a particular exposure scenario.

Regulatory toxicology relies on risk assessment and experimental data to determine the risk and benefits, or the costs and benefits of exposure to certain chemicals, to determine whether such chemical will be allowed in the public sphere and to what extent its use and exposure will be regulated.

Clinical toxicology involves the application of toxicological principles within a diagnostic setting, usually to determine whether a presenting adverse effect or disease or injury is due to some type of chemical exposure.

Forensic toxicology focuses on medical-legal aspects of chemical exposure and toxic injury. Evaluation of analytical chemistry techniques and measurements, and testimony of conclusions and opinions in courts of law encompass the activity of this specialized field of toxicology.

Aquatic toxicology is the study of the effects of manufactured chemicals and other anthropogenic and natural materials and activities on aquatic organisms at various levels of organization, from subcellular through individual organisms to communities and ecosystems.

Chemical toxicology is the study of the adverse effects of chemicals on living organisms.

Ecotoxicology is the integration of toxicology and ecology or, as Chapman (2002) suggested, “ecology in the presence of toxicants”. It aims to quantify the effects of stressors upon natural populations, communities, or ecosystems.

Environmental toxicology and its sister-subject ecotoxicology refers to the scientific study of the potential effects upon organisms of anthropogenic compounds released into the natural environment.

Challenges Wise use of chemicals is an essential component of the

high standard of living we enjoy. The challenge to toxicologists is to ensure that we are not endangering our health or the environment with the products and by-products of modern and comfortable living.

As a career, toxicology provides the excitement of science and research while also contributing to the well-being of current and future generations.

Opportunities With the increase in our health consciousness, as

well as concern for our environment, a wide and growing variety of career opportunities exist in toxicology.

Attractive Salaries and Professional Advancement The demand for well-trained toxicologists continues

to increase. Increasing specialization in the science of toxicology now provides the toxicologist with a competitive advantage over chemists, engineers, biologists or other scientists without specialized training in toxicology.

Research, Product Safety Evaluation, Teaching, Public Service, Regulatory Affairs and Consulting

Where can I find more information? SOT – Society of Toxicology:

http://www.toxicology.org SETAC – Society of Environmental Toxicology

and Chemistry: http://www.setac.org/

Thank you

‘Melaka Education Fair 2009’

Melaka Institute of Biotechnology