Barrick Lab :: ProtocolsDegenerateBases

---+ Use of degenerate bases
Spoke with IDT about some of their recommendations related to incorporation of degenerate bases in oligos.

---++ Machine Mix vs Hand Mix option

   * Machine Mix:
      1 Bases are mixed at equal molar concentrations.
      1 Each base has a different coupling efficiency which leads to different incorporation rates.
         * Automated message states that this is a difference in delivery rates, but that appears to be somewhat misleading.
      1 Results in the following incorporation ratio for a single "N":
         * 30:26:24:20 T>G>C>A
      1 Ratio is per base, and a string of N's does not change the ratio.
      1 No additional charge.
   * Hand Mix:
      1 Can be used to correct the machine mix ratio.
      1 Percents entered are the incorporation percents, not the mixture percents.
         * ie: entering 25:25:25:25 would result in them decreasing the amount of T and increasing the amount of A to achieve 25:25:25:25 final diversity.
      1 Claim their formula is accurate to within less than 1%.
      1 Additional cost required.

---++ PAGE and HPLC problems

   1 Each base (A,C,T,G) has a slightly different migration rate than each other. 
   1 IDT uses a very conservative selection to ensure that all product is full length.
   1 Results in loss of diversity.
   1 Works against hand mixing.
   1 Suggested that can try standard desalting and doing our own less conservative PAGE purification to eliminate truncated product. Warning that this can still lead to elimination of some diversity.


-- Main.DanielDeatherage - 22 Jan 2013
This topic: Lab > WebHome > ProtocolList > ProtocolsDegenerateBases
Topic revision: r1 - 2013-01-22 - DanielDeatherage