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#Bioinformatics  ##Command #Background  ##Background: Bioinformatics  ###Command  Line/Terminal Tutorial This workflow is written assuming that the user is using a computer running Mac OS or Linux. It is also possible to carry out many of the computational parts of this workflow in a Windows environment but getting these steps to work in Windows is outside the scope of this project.   Some parts of this workflow require the user to provide text instructions for software programs by using a command line interface. While potentially intimidating to computer novices, the use of command line interfaces is sometimes necessary (_e.g._, some programs do not have graphical interfaces) and is also sometimes much more efficient. To access the command line on a Mac open the Terminal program (the default location for this program is in the "Utilities" folder under "Applications"). 
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And this interactive tutorial: [http://www.ee.surrey.ac.uk/Teaching/Unix/](http://www.ee.surrey.ac.uk/Teaching/Unix/)  ##Summary ###Summary  of Unix/Linux commands and terms $ **ls** lists files and directories (folders). If left as just "ls" this command will list the files and directories in your current location. If a "path" is added afterwards (e.g., ls /usr) this command will list the files and directories in that location.  $ **cd** use to change directories 
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_compile_ - turning a human-readable file into a computer-executable program  ##Software ###Software  updates Software packages are updated with varying frequencies. Some such updates will render the instructions offered here obsolete. When this occurs, you should consult with the software manual for help. An internet search with a description of the problem you are having may prove helpful. Another option is to email the software developer; many are remarkably responsive. As a last resort, consult with a colleague who is more comfortable with bioinformatics or computer programming. Most software updates will require only minor modifications. For example, we might provide you with instructions to type:  ./software_1.2.0/software.py  but a more recent release might necessitate:  ./software_1.3.0/software.py ##Background: Molecular biology and microbiology  This workflow assumes a basic knowledge of molecular biology and sterile technique (methods for carrying out lab experiments without contamination from living microorganisms). The starting point is the collection of microbes from a surface with a swab. We will cover the steps necessary to take a sample through plating, dilution streaking, overnight growth, creating a glycerol stock, 16S rDNA PCR, and preparation for Sanger sequencing to determine the identity of your bacterial or archaeal isolate.   Throughout the "Isolation" section we refer frequently to "media" and "culture media". This is in reference to the type of substrate (sometimes liquid, sometimes a gel like material such as agar) used to grow microbes in the lab. The choice of media will depend on the goals of the particular project. Some factors to consider when selecting media and conditions for growth include:   1. What type of organism do you want to isolate?  2. Are there types of organisms (_e.g._, pathogens) that you would prefer not to isolate? For example, swabbing people and growing samples on blood agar at 37 °C can preferentially isolate human pathogens.    3. How much time is available for growth and isolation?  + growth rates differ both between organisms (_e.g._, species 1 versus species 2) and also in different conditions for the same organisms (_e.g._, species 1 at 20 °C vs. 37 °C)  + for many microbes there is an "optimal growth temperature" (OGT - the temperature at which it grows best) but the OGT varies between species  + you will be able to isolate a greater diversity of organisms if you allow a long time for slow-growing organisms to grow    4. What types of equipment are available to you?  + if an organism grows most happily at 37 °C, then you will need to have an incubator and shaker available at that temperature.  For our previous work we have simply used a rich media such as lysogeny broth (LB) and growth at either room temperature (~25 °C) or 37 °C. For some basic information on media preparation and agar plates, we recommend the following resource: [http://teach.genetics.utah.edu/content/gsl/html/agar.html](http://teach.genetics.utah.edu/content/gsl/html/agar.html)