Manuscrito científico 2011

National Genomic Research Initiative

Science Education Alliance, Howard Hughes Medical Institute

Department of Biology

University of Puerto Rico at Cayey













Purification and Characterization of Mycobacteriophage Epéneto Isolated From Tropical Soil of Puerto Rico and Bioinformatics Annotation of Mycobacteriophage Cuco Nucleotides 18,647 to 25,743.Authors: Joannieliz Rodríguez Cora, and Michael Rubin, National Genome Research Initiative, Science Education Alliance, Howard Hughes Medical Institute and Department of Biology, University of Puerto Rico at Cayey.

Abstract

The experiment is carried out, consisted in isolating ground from somewhere in our country, Puerto Rico. The purpose of this was to find avirus or as it commonly called: phages. Then, extract its DNA and investigate their characteristics, unique and innovative in the scientific world. The investigations began with isolating a soil sample and insert it into a 15 ml conical tube. Only use two grams of sample to proceed with enrichment, which proposes Increase the likelihood of the phage, obtaining a novel phage from the environment. Weuse a specific host or bacteria to the phage, called Mycobacterium Smegmatis. Then, use aseptic technique to minimize the mycobacterium or microorganisms found in the work area. At the end of the enrichment process, we turn to making dilutions, about three rounds, to find the approximate amount of phage and characterizing them according to their appearance, size and shape. After at least 3 roundsof purification, it proceeds to the "medium titer lysate" for a "web pattern" and even closer to the concentration of phages. Later, we proceed to do the "high titer lysate" or planting of ten plates, to obtain the final concentration and move to remove the genomic DNA. After this last process, the genomic DNA, if the phage is a good candidate, can pass sequenced or saved for future research.After all the hard work in the First Semester of the course, Laboratory of Biology, there were like 30 identified phages to add in the bank of Phages Database. All of the phages pass for many obstacles, but good experiences. These phages were unique and different from the others. Many of them achieve the goal (the genomic DNA), but others only arrived to the Empiric Test, Medium TiterLysate or High Titer Lysate and this was very positive for the investigator and the professor. In the same way, some of the owners of the phages, continued in the Second Semester and they finish the goal. In the last days of the First Semester the group chooses one Phage to be sequenced and that was CUCO, from Valerie Nazario. The Second Semester course of Bioinformatics, we received CUCO sequenced andstart working with the sequence. In this part of the work the group and the professor are going to use some technology tools from the Internet to obtain all the characteristics of CUCO, his cluster, his function and other important things.

Introduction

Bacteriophages (phages) are viruses that infect bacteria. Typical phages have hollow heads (where the phage DNA or RNA isstored) and tunnel tails, the tips of which have the ability to bind to specific molecules on the surface of their target bacteria. The viral DNA is then injected through the tail into the host cell, where it directs the production of progeny phages often over a hundred in half an hour. These "young" phages burst from the host cell (killing it) and infect more bacteria. Bacteriophages are the mostabundant forms of life in the biosphere and carry genomes characterized by high genetic diversity and mosaic architectures.

The genomic analysis of 30 complete mycobacteriophages—viruses that infect mycobacterial hosts—reveals them to be genetically diverse and to contain many previously unidentified genes. The abundance and diversity of phages, the simplicity of phage isolation, and the...