Semester 4-Week 6-This Semester's Research Proposal

Hello! Welcome back. Since we have research proposals due this week, I am including mine here as my blog post. Please note that as my project is wrapping up, this submission does not include the normal hypothesis/projected results format that a proposal normally would. Instead, this is a preliminary version of my project abstract. Thanks for checking in this week-I appreciate your patronage!

Abstract

Since the complete genetic sequencing of the Escherichia coli genome in 1997 (Blattner, et.al), variations of the 16s ribosomal gene have been found conserved among different bacterial species. As a result, 16s ribosomal DNA sequencing has become an integral part of bacterial identification in the modern laboratory (Janda & Abbot, 2007). The initial step of the identification process requires extraction of DNA, which must then be amplified using a polymerase chain reaction (PCR) so it can be sequenced. Primers specific to certain regions of the 16s gene are utilized to replicate the bacterial DNA during the PCR process so that the resultant DNA can be sequenced and correctly identified (Mao et. al 2012). Because PCR amplification is in widespread laboratory use, knowledge of DNA extraction and PCR techniques is an essential component of the student learner’s skill set. This study identified and designed simple laboratory protocols for use in introducing the Phoenix College student learner to elementary DNA identification technology. The protocols are intended to accompany the standard clinical format of culturing both known and unknown bacteria. The study identified electrophoresis gel techniques, extraction protocols for gram-negative and gram positive bacterial species, a set of universal 16s ribosomal primers that can be used to identify fifteen gram-negative and gram-positive bacterial species available in the Phoenix College Biosciences laboratory, and a congruent PCR amplification protocol that can be adapted and incorporated into existing instructor curricula.

References:

References: Blattner, R., Plunkett III, G., Bloch, C., Perna, N., Burland, V., Riley, M., Collado-Vides, J., & Glasner, J. (1997). The complete genome sequence of escherichia coli k-12. Science, 277, 1453-1462. doi: 10.1126/science.277.5331.1453.

Cattelino, P. (2014) Identification and application of universal 16s rRNA ribosomal primers. Phoenix College Student Paper. Pps. 1-16.

Frank, J., Reich, C., Sharma, S., Weisbaum, J., Wilson, B., & Olsen, G. (2008). Critical evaluation of two primers commonly used for amplification of bacterial 16s rRNA genes. 74(8), 2461-2470.

Janda, J., & Abbot, S. (2007). 16s rRNA gene sequencing for bacterial identification in the diagnostic laboratory: Pluses, perils, and pitfalls. Journal of Clinical Microbiology, 45(9), 2761-2764.

Mao, D., Zhou, Q., Chen, C., & Quan, Z. (2012). Coverage evaluation of universal bacterial primers using the metagenomic datasets. BMC Microbiology, 12(66), Retrieved from www.biomedcentral.com/1471-2180/12/66.

Marchesi, J., Soto, T., Weightman, A., Martin, T., Fry, J., Hiom, S., & Wade, W. (1997). Design and evaluation of useful bacterium-specific PCR primers that amplify genes coding for bacterial 16s rRNA. Applied and Environmental Microbiology, 64(2), 795-799.

PS: Just because I LOVE space exploration and NASA, here is a link to NASA's news page and an image of one of Jupiter's moons, Io, for your enjoyment.

This global view of Jupiter’s moon, Io, was obtained during the tenth orbit of Jupiter by NASA’s Galileo spacecraft. Credit: NASA