From Dirt to Streptomyces DNA

The purpose of this semester-long Lesson is to give students an authentic, course-based undergraduate research experience during which they learn basic and advanced microbiological and molecular biology techniques. This project begins with the isolation of a suspected Streptomyces bacterium from a soil sample and concludes with its identification. Students collect data, regarding colony and cell morphology, biochemical characteristics, the production of secondary metabolites, and employs the PCR using custom-designed primers to the Streptomyces 16s rRNA gene. The project culminates with the identification of their soil isolate using the National Center for Biotechnology Information (NCBI) web site to perform nucleotide blasts. The blastn program provides the final piece of evidence used to confirm, or not, the identification of their isolate as a Streptomyces from 16s rRNA gene sequence data, hence the title “From Dirt to Streptomyces DNA. In addition, the Lesson focuses on the Streptomyces bacteria to address several ASM aligned goals and objectives. These include prokaryotic growth phases and ways in which interactions of microorganisms among themselves and with their environment is determined by their metabolic abilities. In addition, this Lesson illustrates how microbial metabolism is important to a relevant societal issue, the need for new antibiotic discovery particularly given the rise of antibiotic resistance strains of clinically relevant bacteria. It also illustrates the microbial diversity of soil and the developmental/physiological strategies employed in such a competitive environment. This Lesson hopes to impart both the thrill and challenges associated with scientific discovery. Citation: Brodkin MA. 2021. From dirt to Streptomyces DNA. CourseSource. https://doi.org/10.24918/cs.2021. 32 Editor: Rachel Horak, American Society for Microbiology Received: 1/9/2020; Accepted: 4/30/2021; Published: 10/25/2021 Copyright: © 2021 Brodkin. This is an open-access article distributed under the terms of the Creative Commons Attribution-NonCommercial-ShareAlike 4.0 International License, which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original author and source are credited. Bio-Protocol LLC has given written permission via email for protocol use in LAB 8 (S7). Qiagen has granted permission to use their protocol in LAB 8 (S7) with a written Image Transfer Agreement. Genewiz.com has given written permission via email to use screen shots from their web site in LAB 11(S10). NCBI NBLAST site is an open-source web site. Conflict of Interest and Funding Statement: The Widener University Provost Grant Program supported part of this work. The author does not have a financial, personal, or professional conflict of interest related to this work. Supporting Materials: Supporting Files S1. Dirt to DNA-Handout Introduction to the Actinobacteria (LAB 1); S2. Dirt to DNA-Handout Serial dilution and spread plate technique (LAB 2); S3. Dirt to DNA-Handout Count serial dilution plates and subculture to ISP medium 2 (LAB 3); S4. Dirt to DNA-Handout slide culture technique (LAB 5); S5. Dirt to DNA-Handout Check pure culture isolates for antibiotic action (LAB 6); S6. Dirt to DNA-Handout Grow suspected Streptomyces isolates in broth culture (LAB 7); S7. Dirt to DNA-Handout DNA extraction from broth culture of Streptomyces isolate (LAB 8); S8. Dirt to DNA-Handout Agarose gel electrophoresis of DNA extraction and UV scan (LAB 9); S9. Dirt to DNA-Handout Polymerase Chain Reaction (LAB 10); S10. Dirt to DNA-Handout PCR Product Sequencing of 16s rRNA gene at Genewiz.com (LAB 11); S11. Dirt to DNA-Handout *NCBI Nucleotide Blast (blastn) (LAB 12); S12. Dirt to DNA-Handout Laboratory Report Outline (LAB 13); S13. Dirt to DNA-Bacterial cultures, Media, Solution recipes, and Equipment; S14. Dirt to DNA-DNA extraction protocols instructor copy; and S15. Dirt to DNA-Student Supporting Files LAB 1-12. *Correspondence to: Marc Brodkin. Widener University, Department of Biology, One University Place, Chester, PA 19013. Email: brodkinmarc22616@gmail.com CourseSource | www.coursesource. org 2021 | Volume 08 1 Lesson