1. Biology Project Overview Comparisons of GAPC Gene Organization in Plants

2. GAPC Gene Codes for the enzyme: Glyceraldehyde-3-Phosphate Dehydrogenase (Cytosolic)Codes for the enzyme: Glyceraldehyde-3-Phosphate Dehydrogenase (Cytosolic) Gene is found in the cell nucleusGene is found in the cell nucleus Member of a Gene FamilyMember of a Gene Family –4 genes in the thale cress plant Arabidopsis, a member of the mustard plant family –GAPC and GAPC-2 code for enzymes that act in the cell cytoplasm –GAPCP-1 and GAPCP-2 code for enzymes that act in the chloroplasts

3. GAPC Gene Product Enzyme (gene product) works in the cell cytoplasmEnzyme (gene product) works in the cell cytoplasm Reaction:Reaction: NADH is an electron carrier that donates electrons to the Mitochondrial Electron Transport Chain for production of ATPNADH is an electron carrier that donates electrons to the Mitochondrial Electron Transport Chain for production of ATP ATP (Adenosine Triphosphate) provides energy for most endergonic biological reactionsATP (Adenosine Triphosphate) provides energy for most endergonic biological reactions NAD + (Nicotine Adenine Dinucleotide) accepts one proton and two electrons to become NADH Glyceraldehyde-3- Phosphate Dehydrogenase is the catalyst

4. Using Polymerase Chain Reaction (PCR) PCR is a highly accurate and efficient way of copying (amplifying) a specific region of DNA for studyPCR is a highly accurate and efficient way of copying (amplifying) a specific region of DNA for study The process involves specific primers that attach to the end of the target sequence and serve as starting points for elongationThe process involves specific primers that attach to the end of the target sequence and serve as starting points for elongation Heating and cooling cycles provide multiple rounds of PCR where the template number doubles with each roundHeating and cooling cycles provide multiple rounds of PCR where the template number doubles with each round Taq polymerase, a heat-stable enzyme that catalyzes DNA synthesis, is used to produce the DNA copiesTaq polymerase, a heat-stable enzyme that catalyzes DNA synthesis, is used to produce the DNA copies

5. Using Polymerase Chain Reaction (PCR) Starting with a single copy of a DNA sequence, over 1 million copies can be generated with about 30 rounds of PCRStarting with a single copy of a DNA sequence, over 1 million copies can be generated with about 30 rounds of PCR In the Biology Research Project, students will amplify the nuclear gene for glyceraldehyde phosphate dehydrogenase (GAPC) from a variety of plants, with a “nested” procedure using two sequential sets of primers to enhance specificityIn the Biology Research Project, students will amplify the nuclear gene for glyceraldehyde phosphate dehydrogenase (GAPC) from a variety of plants, with a “nested” procedure using two sequential sets of primers to enhance specificity

6. Using Polymerase Chain Reaction (PCR) Inner Arrows show position of primers for PCR Round 2 Outer Arrows show position of primers for PCR Round 1 Sizes of DNA Products can be determined by agarose gel electrophoresis GAPC gene PCR product is missing one intron (interruption) found in the other three genes

7. Comparing Round 1 and Round 2 Results Size Markers Negative Control Positive Control (genomic) ParsleyOregano Positive Control (plasmid) ROUND 1 Size Markers Negative Control Positive Control (genomic) ParsleyOregano Positive Control (plasmid) ROUND 2

8. Join the Biology Research Project if You: Are interested in applying molecular biology techniquesAre interested in applying molecular biology techniques –Polymerase Chain Reaction –Agarose Gel Electrophoresis Would like to explore evolutionary relationshipsWould like to explore evolutionary relationships –Sizing DNA Fragments –Bioinformatics of Sequence Data