Prem Chourey, Mentor
Ranyelle Craig is preparing the buffer used to rupture the maize cells from which nucleic acids are extracted.
Preparing agarose gel, which will be used in resolving mixtures of DNA fragments.
Preparing a sample for the spectrophotometer.
Genetic Cloning: A Scientific Process
The main objective of my research at the Crop Genetics & Environmental Research Unit, CMAVE, Gainesville, Florida, was to gain hands-on training and experience with a few essential techniques that are routinely applied in Plant Molecular Biology research. More specifically, I isolated deoxyribonucleic acid (DNA) from Escherichia coli (E. coli) and genomic DNA from etiolated Zea mays shoots. I also isolated ribonucleic acid (RNA) from maize suspension-cultured cells. I was able to quantitate the nucleic acids using a spectrophotometer with an absorbency of 260 AU. I used agarose gel electrophoresis to assess the quality of the nucleic acids. The RNA extracted was used to perform a reverse transcription, which uses an RNA template to generate single-stranded DNA molecules complementary to RNA (cDNA). Subsequently, a polymerase chain reaction was applied, amplifying the cDNA exponentially through cycles of denaturation, annealing and extension. This method, more commonly referred to as RT-PCR, allowed the successful partial cloning of a Sucrose Synthase gene.