This experiment aims to isolate and sequence the GAPDH gene from the Cucumis sativus plant. GAPDH is an important enzyme that catalyzes a step in glycolysis. The experiment involves extracting DNA from C. sativus, amplifying the GAPDH gene via PCR, separating the DNA fragments via electrophoresis, purifying the DNA, and attempting to insert the gene into bacteria for replication and sequencing. However, the experiment was not completed because the bacteria needed for DNA purification did not grow, likely due to the presence of an antibiotic.

1. By: Erick G. Rodríguez & Jefry Lopez

2.  The main purpose of this experiment is to isolate and
sequence the GAPDH (glyceraldehyde-3-phosphate
dehydrogenase) gene from the Cucumis sativus plant.
 GAPDH
 An enzyme
 Catalyzes the sixth step of glycolysis
 Serves to break down glucose for energy and carbon molecules.
 Most abundant enzyme in cells

3.  Dicot
 Grows up by wrapping
around objects using
spiraling tendrils.
 Its fruits are called
cucumbers.

4.  Kingdom: Plantae
Division: Magnoliophyta
Class: Magnoliopsida
Order: Cucurbitales
Family: Cucurbitaceae
Genus: Cucumis
Species: C. sativus

7.  Nucleic Acid Extraction
 Separate DNA from other “waste components” of the cell.
 GAPDH PCR
 Amplify a specific segment of the GAPC gene by utilizing repeated
cycles of DNA synthesis.
 Electrophoresis
 Separate DNA fragments by size using an electric current that
passes through the agarose gel.

8.  PCR Purification
 To clean DNA from all the impurities that result from the PCR
using Chromatography.
 Ligation
 To join two piece of linear DNA to form one single piece.
 Catalyzed by an enzyme called DNA ligase.
 Transformation
 Used to introduce the plasmid into living bacterial cells so it can be
replicated

9.  Plasmid purification
 Perform a miniprep of the plasmid DNA in preparation
for DNA sequencing.

• DNA Sequencing
 Determine the exact order of nucleotide bases.
• Compare our results with the other data bases

11. Future steps
Plasmid purification
 Perform a miniprep of the plasmid DNA in preparation for
DNA sequencing.
•DNA Sequencing
Determine the exact order of nucleotide bases.
•Bioinformatics
Compare our results with the other data bases

12. Conclusion
This experiment was not completed because the bacteria
needed for the DNA purification didn’t grew. One of the possible
explanations for it is that because of the Amphiciline this
bacteria didn’t grew. So the next steps couldn’t be completed.