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Secugen - Technical Solutions - Stuttering After Mononucleotide Stretches

Sequence Appearance.

  • Sequencing data quality is poor after stretches of 10 or more nucleotides of the same base due to mixed bases.

Example.

Stuttering

Stuttering After Mononucleotide Stretches

Possibles Causes.

  • Polymerase slippage during DNA synthesis. This is a recognized limitation of the Sanger method.

Treatment.

  • Sequence from the reverse direction.
  • Use a poly-mononucleotide primer with a degenerate base (wobble) at the 3’ end.