I would like to ad a strategy about how many blue and green nucleotides I will allow in line right after each other on a strand of a certain length.
This is because green and blue nucleotides seem to be working as stabilizers, if there are not too many in line, compared to the length of string they appear on. If too many, they will destabilize the design. But in the right number, I think they are stabilizors.
So here is my numbers:
In a string 3 nt long, allow max 3 blue/green nt in line on a strand
In a string 4 nt long, allow max 3 blue/green nt in line on a strand
In a string 5 nt long, max 3
In a string 6 nt long, max 4
In a string 7 nt long, max 4
In a string 8 nt long, max 4
In a string 9 nt long, max 4
In neckarea up to 6 is allowed. (including this number)
Penalize with -1 for each extra blue/green nucleotide above the number allowed