Exploring Clustered Regularly Interspaced Short Palindromic Repeats Diversity in Soil Bacteria

Abstract

Clustered regularly interspaced short palindromic repeats (CRISPR) is a defense system found in bacteria (and archaea) against bacteriophages, viruses that infect bacteria. CRISPR DNA sequence consists of short, identical nucleotide repeats that are interspaced by spacers. The spacer sequences are records of viral infections experienced by the bacteria and play the role of an “immunization card”: if the spacer sequences match the infecting viral DNA, the bacteria will cut the viral DNA with CRISPR‐associated (Cas) proteins, eliminating the threat of viral infection. We speculate that the spacer sequences reflect the history of bacteriophage infections, which are unique to the environment bacteria reside in. Environmental factors, such as location and time, weather, local ecosystems, and pollutions, will alter the spacer sequences found in bacteria. This research focuses on investigating and discovering new spacer sequences from soil samples collected around NY/NJ metropolitan area, a historically heavily polluted urban ecosystem, at various locations, timepoints, and weather conditions. To accomplish this goal, the genomic DNA was extracted and purified from the collected soil samples. The CRISPR locus was amplified by using polymerase chain reaction (PCR) and then purified via gel electrophoresis. The purified CRISPR DNA was then ligated into a p‐Drive vector and transformed into M15 competent cells. The cells were cultured to amply the vector containing ligated CRISPR sequence, which was subsequently analyzed by aligning against CRISPR spacer database to identify new space sequence. We expect that the newly discovered spacer sequences reflect the history of viral infections unique to the regions investigated, which can potentially inform the understanding of health risks related to environmental bacteria and viruses.Support or Funding Information‐ U.S. Education Department Minority Science & Engineering Improvement Program grant # P120A160084‐ NIH AREA Grant # CA179410‐ USED Title III Part F, HSI‐STEM & Articulation Grant #P03C160155This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.