“Hand-Mixed Bases”, also known as degenerate, random, or wobble bases, can be introduced at any position in an oligonucleotide sequence. We use a different notation for Hand-Mixed Bases than for our standard machine mixed bases. Thus, when inputting your sequence, you simply choose the mixed base letter (IUB symbol) that represents the desired base mix from the table below.
For the first occurrence of a particular mixed base, the proportion of each base must be indicated, in the order ACGT. For example, an N mixed base that requires 25% of each included base will be indicated as (N:25252525); if the bases are required in the proportions 26% of A and T, and 24% of C and G, the mixed base will be represented as (N:26242426). Each mixed base after the first insertion of that ratio will not need the ratio, just the base in parentheses, e.g. (N).
Multiple mixes are possible, for a total of 4 maximum per oligo.
See example sequences below:
Example 1: ACGSSSACT with 50% of C and G for all of the S bases would be indicated as follows: ACG(S:00505000)(S)(S)ACT
Example 2: ACTGACTGNNNNNNNNNACTGATGC with 25% of A, C, G, and T for all of the N bases would be indicated as follows: ACTGACTG(N:25252525)(N)(N)(N)(N)(N)(N)(N)(N)ACTGATGC
Example 3: ACTGACTGNNNNNNNNNNCAGTACTG with the first 5 N-bases being equimolar, and the latter 5 N-bases being a 70/10/10/10 mix (favoring A): ACTGACTG(N1:25252525)(N1)(N1)(N1)(N1)(N2:70101010)(N2)(N2)(N2)(N2)CAGTACTG
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