PLANTS AS BIOREACTORS

DNA extraction

• Hina Amir

• Shakiba Anwar

• Sadaf Moneeba

DNA extraction (human)

PCR (gene of interest)

Transformation

Tissue Culture

Southern blotting Northern blotting Western blotting

DNA extraction and RFLP

RNA extraction and gel electrophoresis

Protein extraction and PAGE

PRINCIPLE

DNA Isolation

• Removal and separation ofDNA.

1. Cell Lysis

2. Precipitation

3. Wash

4. Resuspension

• human blood

• DNA

TOPICAPPLICATIONS

DNA sequencingCheck genetic diseasesPCR templateAnalyze forensic evidence

PRINCIPLE

Polymerase Chain Reaction

Exponential amplification sequence of interestlinked to replication in vivo

APPLICATIONSTOPIC

• Gene

•

COPIESOF

GENE

Direct cloning from genomic DNA or cDNA

Analyze forensic samples

Increase amount of DNA

PRINCIPLE

Plant Transformation

integration (foreign DNA ) cell regeneration

Expressing gene of interest

APPLICATIONSTOPIC

• Gene of interest

• vector

Disease free plants Improve quality of plants (saltresistance, drought resistant plants)Hormone expressionpest resistance

PRINCIPLE

Tissue Culture

cell or tissue or organ fully differentiated organism

APPLICATIONSTOPIC

• Transformed Tobacco cells

Synthetic hormones AntibodiesArtificial pancreas —islet cellsArtificial bladders Cartilage.Artificial skin

PRINCIPLE

DNA Isolation

TOPIC

• DNA from cell cultured plant

PRINCIPLE

Restriction fragment length polymorphism

APPLICATIONSTOPIC

• Restriction fragments of required length

Paternity cases

Criminal cases

Disease status of an

individual

PRINCIPLE

Southern Blotting

APPLICATIONSTOPIC

Amount of gene or DNA

DNA profiling

Proving someone guilty or innocent

Paternity Testing

PRINCIPLE

RNA Isolation

1. Cells or tissue disruption2. Inactivate RNases3. Denature nucleic acid-protein

complexes4. RNA isolated

TOPIC

• RNA from plant

PRINCIPLE

Gel electrophoresisTOPIC

• Proteins from plant• Migration of charged

molecules in an electric field through a solution or solid support

• Polyacrylamide –Proteins

• Good preparative and analytical method

PRINCIPLE and APPLICATIONS TOPIC

• Northern blottingSample preparing.Transfer.Adding probe. Incubation.Blot visualization

Gene's expression pattern

PRINCIPLE and APPLICATIONS TOPIC

• Proteins Adding SDS.

heat.

Transfer of denature protein.

Loading sample.

SDS-PAGE

PRINCIPLE and APPLICATIONS TOPIC

• Proteins Sample preparation.

Transfer.

Incubation.

Adding antibody.

Confirmatory test in HIV.

REFERENCES

• MOLECULAR BIOLOGY LAB May 2010 Thoraia Shinawi

• Rodríguez-Lázaro, D. and Hernández, M. (2013). Introduction to the Real-time. Real-Time PCR in Food Science: Current Technology and Applications.Caister Academic Press, UK. ISBN: 978-1-908230-15-7.

• http://bioneer.com.au/Products/DNA-RNA-Preparation-Kits

• http://science.howstuffworks.com/dna-profiling1.htm

• http://www.bio.davidson.edu/genomics/method/RFLP.html

• http://www.highveld.com/pcr/principle-of-pcr.html

• http://www.jains.com/Tissue/images/tissue_main.jpg