Applied Biosystems 3730xl DNA Sequencer (operated by Core Staff)

 

ABI 3730XL Sanger sequencer

Applications

  • This instrument is used by core personnel to sequence DNA templates
  • Sequencing of cloned inserts or PCR products is done by DNA polymerase-mediated linear amplification of templates in the presence of mixtures of dNTPs and fluorescently-labeled dideoxynucleotide triphosphates (Sanger method), followed by fractionation of the extension products by capillary electrophoresis using the 3730xXL.
  • Although Sanger sequencing is well established and lower throughput that modern NGS sequencers, it is still the gold standard in terms of accuracy and is still used worldwide for the low cost sequencing of plasmid and PCR templates.
Specifications

  • This is a capillary sequencer capable of processing 96 template samples simultaneously; up to ~4000 samples/week.
  • Standard sequencing PCR reaction conditions are:
  1. 96 degree C denaturation 30 seconds 
  2. 50 degree C extension for 15 seconds  
  3. 60 degree C extension for 4 minutes 30 seconds
  4. A total of 30 cycles are used
  • We use conditions that should work for most templates. Please consult with Genomics Core staff for troublesome templates requiring different conditions.
  • Submit templates in either individual reaction tube format or cost-effective plate format.
  • See Sanger Sequencing Services for directions about sample submission utilizing this instrument.