Lecture/DNA_Barcoding

Analyses for DNA Barcoding †

• Requests from USU, Forestry
  • 2021.03.16

Softwares for Sanger Sequencing Analyses †

• See this web page of Genewiz: https://www.genewiz.com/Public/Resources/Tools-for-Viewing-Sequencing-Data/
  The following three will be good candidates to edit and assemble Sanger Seq files
• UGENE http://ugene.net/
  • Looks Fine!: To be used in this lecture
• GeneStudio Pro http://genestudio.com/genestudio
  • Windows Only. Looks good.
• Chromaseq http://mesquiteproject.org/packages/chromaseq/manual/
  • Looks good, but a bit older?

Data provided by people at USU †

• Sanger sequencing data:

• Sample sheet
  • Fish, Crab, ...Benthos: 54 samples
    • many for 16S
  • Excel sheet or any datasheet is necessary: ■Rahmah Hayati will prepare the sheet.

• Primers
  • 16S for many samples and 12S for four samples
  • Primer sequences and references are necessary
  • Primers: 16S: product ca 550bp

    16S CCTGTTTACAAAAACAT
    16SR: AGATAGAAACCAACCTGG

  • Primers: 12S: product ca 200bp

    12S TTTCATTCCTTGCGGTAC
    12S AAAGCACGGCACTGATGC

• PCR condition
  • Temperature, time, ...
  • PCR cycles: 35 - 40 cycles of

     92 - 58 - 72

sequence data assemble †

• Please install one of the sequence viewers to your PC. (for Mac: 4 peaks, for win: Sequence Scanner 2)
• Alsop, Please install one of the sequence editors from the link above (for Mac: UGENE, for win UGENE or Gene Studio)
• 1. Sequence assemblage by using Forward and Reverse sequences.

• 2. using AliView (https://ormbunkar.se/aliview/), get Reverse Compliment seq and align together.