Team:Dundee/Team -




iGEM Sumbawagen 2014 · Econey

Notebook – Protocol – 10. DNA Sequencing

DNA sequencing was done by ordering to commercial company.

1. Premix sample was prepared with the following composition:

  • Plasmid sample, 10 to 20 ul*
  • 10 uM primer, 1 ul
  • Sterilized distilled water, 9 to 19 ul
  • Total volume, 21 ul
    Concentration < 900 ng

2. Samples were sent to the company

3. According to the company, DNA sequencing was done using the following conditions:

  • Sequencing kit, BigDye® Terminator v3.1 Cycle Sequencing Kit (Applied Biosystems)
  • Purification kit, BigDye XTerminator® Purification Kit (Applied Biosystems)
  • Sequencer, Applied Biosystems 3730xl DNA Analyzer

4. DNA sequencing data in .ab1 files were received by email, then analyzed using Bioedit program. For alignment, online CLUSTAL program was used.