2009-ag-1266 Msc.(hons.) Agricultural Biotechnology ...icdd.uaf.edu.pk/Presentations/001.pdf ·...
Transcript of 2009-ag-1266 Msc.(hons.) Agricultural Biotechnology ...icdd.uaf.edu.pk/Presentations/001.pdf ·...
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Muhammad Amjad Nawaz2009-ag-1266
Msc (hons ) Agricultural BiotechnologyMsc.(hons.) Agricultural BiotechnologySupervised by
Dr. Iqrar Ahmad Khan
Center of Agricultural Biochemistry and Biotechnology (CABB)UNIVERSITY OF AGRICULTURE
FAISALABAD
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Contents
Genus Aspergillus
Aspergillus niger
Glucose oxidase (Gox)
Molecular markersMolecular markers
Objectives of the study
Progress
Planning ahead
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Group of filamentous fungiGroup of filamentous fungi
180 species180 species
Causes diseasesCauses diseases
Economically importantEconomically important
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H l id
1• Haploid • Filamentous fungi
C t ffi i t t t l ti l difi ti
2
• Cary out efficient post translational modification• Produces enzymes, afflatoxins, mycotoxins,
PGRs2
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RAPD, SSR, RFLP &
Brilliant tool
RAPD, SSR, RFLP & transposons are mostly
used
DNA markers
Helps to understand Measures geneticpolymorphism in mutants
Measures genetic distances
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objectivesjEvaluation of genetic distance among 20 mutants and wild
type strain of A.niger UAF
Estimating the level of transcription of GOx gene.
l f f d h d f hExploitation of information generated in the studies for the stable and enhanced GOx production
Exploitation of results to prepare Glucose diagnostic kitExploitation of results to prepare Glucose diagnostic kit.
Isolation and cloning of GOx gene with high transcription level in microbial species.level in microbial species.
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Progress
20 mutants and parent strain of A. niger have b b i d f E Bi h lbeen obtained from Enzyme Biotechnology lab. Department of Chemistry and Biochemistry University of AgricultureBiochemistry, University of Agriculture, Faisalabad.
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Methodology used
Growth of Mutants on Potato Dextrose Agar
Methodology used
Growth of Mutants on Potato Dextrose Agar
(PDA)a- Medium preparation and composition
b- Streaking and culture conditions:
Production of Mutants on Vogel’s Mediuma- Medium preparation and compositionp p p
b- Inoculation and culture conditions
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Media composition
Potato Dextrose Agar Vogel’s medium
Ch i l Q iChemical Quantity (g/100ml)
Glucose 2g
Chemical Quantity (g/100ml)
Glucose 2g
Agar 2g
Starch 2g
Peptone 0.1g
MgSO4.7H2O 0.02g
Urea 3g
KH2PO4, 0.008g
NH4NO3 0.2g
(NH4)2 SO4 0.4g
MgSO4.7H2O 0.05g
ZnSO4.7 H O
0.001g
Tri-sodium citrate
0.5g
Yeast 0.2gH2OKCl 0.015g
extractg
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Planning ahead
Optimization of protocol for extraction of DNA of A. niger
Molecular characterization of A. niger UAF mutants by RAPD and SSR
Optimization of protocol for extraction of total RNA andOptimization of protocol for extraction of total RNA and mRNA (Gox gene)
Development of cDNA from mRNA of Gox gene
Determining level of transcription of Gox gene
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THANKSTHANKS