Seqanswers Leaderboard Ad

Collapse

Announcement

Collapse
No announcement yet.
X
 
  • Filter
  • Time
  • Show
Clear All
new posts

  • Demultiplexing combinatorial barcodes from paired end data

    Hey, I've been trying to figure out how to demultiplex paired-end 250 miseq data that has three different barcodes.

    The samples all come from 96 well plates. I have two 5' barcodes representing plate number as well as plate column. These two barcodes are linked via a common adapter sequence. I then have one 3' barcode that represents the plate column.

    I am fairly sure I would be able to demultiplexing the data in 2 rounds in which I first demultiplex based upon each plate/column combination, and then demultiplex the result based upon the 3' barcode.

    This seems fairly cumbersome as I have 2880 unique samples. Is there some sort of software in which I can provide simply the three unique barcodes, and then demultiplex in one step?

    Also, I usually use the A5Pipeline (https://code.google.com/p/ngopt/wiki/A5PipelineREADME) to error correct sequence reads, remove Illumina adapter sequences, and assemble the paired-reads. Would I be able to assemble the reads using A5 and then demultiplex the data, or do I have to run the pipeline on each individual sample after first demultiplexing. Thanks!

  • #2
    Does anyone have experience with combinatorial barcodes?

    Comment


    • #3
      How long are these barcodes? It may be possible to write some custom code to do this in one step but without seeing what you reads look like (or a good schematic) no one is going to be able to help. It may be best to find someone locally who can work with you interactively.

      If the A5Pipeline changes the context of the barcodes it will make things complicated. You may have to run the pipeline post demultiplexing.

      Comment


      • #4
        This will do it https://code.google.com/p/jmhc/

        Comment


        • #5
          Originally posted by JackieBadger View Post
          Can you import fastq files? Description states that it operates on fasta files (internally?). I don't think this will do 3 barcodes in a single pass.

          Comment


          • #6
            use skewer

            Hi ddelalca,
            You may use skewer for this purpose. An example command line is as follows:
            Code:
            skewer -x V3-forward.txt -y V4-reverse.txt --mode ap -t 4 ampliconReads-pair1.fastq ampliconReads-pair2.fastq assigned/
            If you only use some of the combinations, you may specify the matrix by -M as well.
            Good luck!
            Attached Files

            Comment

            Latest Articles

            Collapse

            • seqadmin
              The Impact of AI in Genomic Medicine
              by seqadmin



              Artificial intelligence (AI) has evolved from a futuristic vision to a mainstream technology, highlighted by the introduction of tools like OpenAI's ChatGPT and Google's Gemini. In recent years, AI has become increasingly integrated into the field of genomics. This integration has enabled new scientific discoveries while simultaneously raising important ethical questions1. Interviews with two researchers at the center of this intersection provide insightful perspectives into...
              Yesterday, 02:07 PM
            • seqadmin
              Multiomics Techniques Advancing Disease Research
              by seqadmin


              New and advanced multiomics tools and technologies have opened new avenues of research and markedly enhanced various disciplines such as disease research and precision medicine1. The practice of merging diverse data from various ‘omes increasingly provides a more holistic understanding of biological systems. As Maddison Masaeli, Co-Founder and CEO at Deepcell, aptly noted, “You can't explain biology in its complex form with one modality.”

              A major leap in the field has
              ...
              02-08-2024, 06:33 AM

            ad_right_rmr

            Collapse

            News

            Collapse

            Topics Statistics Last Post
            Started by seqadmin, 02-23-2024, 04:11 PM
            0 responses
            45 views
            0 likes
            Last Post seqadmin  
            Started by seqadmin, 02-21-2024, 08:52 AM
            0 responses
            61 views
            0 likes
            Last Post seqadmin  
            Started by seqadmin, 02-20-2024, 08:57 AM
            0 responses
            51 views
            0 likes
            Last Post seqadmin  
            Started by seqadmin, 02-14-2024, 09:19 AM
            0 responses
            65 views
            0 likes
            Last Post seqadmin  
            Working...
            X