deletions | additions       diff --git a/16S rDNA Sequencing and Analysis (Organism Identification).md b/16S rDNA Sequencing and Analysis (Organism Identification).md  index de1c3e5..f12b2ec 100644  --- a/16S rDNA Sequencing and Analysis (Organism Identification).md  +++ b/16S rDNA Sequencing and Analysis (Organism Identification).md 
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This pipeline allows you to upload one zipped folder containing multiple .abi traces. It cleans and processes the sequences and generates a FASTA file of the processed sequences; which can then be uploaded to BLAST and analyzed. This allows you to quickly screen your samples before running the files through the more time consuming SeqTrace analysis which will reverse complement and align the reads to generate a consensus sequence.  After signing in to RDP you will be on the Library Run Summary page. Click on the Create New Run tab near the top of the page. Select the appropriate 16S rRNA gene (Archaea or Bacteria depending on your sample) name your library and choose a library name abbreviation and select any vector (this pipeline assumes cloned PCR fragments but will work fine regardless of what you select here). Select the Upload "Upload  the data without well mapping mapping"  button at the bottom of the page. You will now be directed to the Data Loader page, choose a zipped folder containing the abi traces you wish to analyze and click Load Data (to create the folder, put all of the abi traces you are working with into a folder, right click on the folder and select Compress “folder name”—if you downloaded the files as a group from your sequencing facility they may already be in a zipped folder). When the pipeline is finished, you will be directed to click a link that will open a new window containing the library run stats. Select the Download Raw Sequence button. Navigate to [http://blast.ncbi.nlm.nih.gov/Blast.cgi?PROGRAM=blastn&PAGE_TYPE=BlastSearch&LINK_LOC=blasthome](http://blast.ncbi.nlm.nih.gov/Blast.cgi?PROGRAM=blastn&PAGE_TYPE=BlastSearch&LINK_LOC=blasthome) and select the Choose File button underneath the area for the FASTA sequence. Select the file you just downloaded from the library run stats page. We recommend checking the box to exclude Uncultured/environmental sample sequences then click BLAST. If you are working with a large number of FASTA sequences it may take a few minutes. When the BLAST search is complete, you can cycle through the sequences you blasted using the pull down menu to the right of the Results for: heading.  
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[http://www.python.org/downloads/](http://www.python.org/downloads/)  After downloading and unpacking the program, SeqTrace is ready to go. for use.  SeqTrace must be launched from a Terminal window. For a refresher or introduction to the Terminal, see section 2. Move SeqTrace to your Applications folder. Open a Terminal window and copy/paste or type: