Consequences of partial CDS optimization

[i-hate-this-username]

Hi there,

If this isn't the right place to ask, please let me know!

In our lab, we're utilizing a library of modules for the N and C terminals because they are repetitive for our experiments. This helps us save on the everyday generation of oligos. Consequently, we don't typically need to optimize the entire coding sequence; we focus primarily on the central part, keeping the terminal sections (50-100 bp) unchanged, and therefore "finalize" the plasmid and the whole protein sequence during the Golden Gate assembly.

Here's my question: When we use the "AvoidChanges" constraint for these terminal parts that are the parts of the protein sequence, could it potentially impact the optimization process?

For example, suppose we have a coding sequence of 1000 bp (positioned from 0 to 1000) with constraints to avoid changes from positions 0 to 100 and 900 to 1000. Should we consider adding any other parameters specific to this kind of optimization constraint?

Thanks!

[veghp]

Hi,
AvoidChanges should be sufficient in itself, shortly, as a constraint it will keep the sequence or fail. Note it can be used as an objective too, but that does not seem to be your use-case.

A specification can be either a hard constraint, which must be satisfied in the final sequence, or an optimization objective, whose score must be maximized.

As you want to preserve the sequence as is, AvoidChanges alone seems to be sufficient, and can define other constraints and objectives on the rest of the sequence. Note that you must use enforcetranslation on the optimised section to ensure it keeps the aa sequence, and that spec must be defined on a length that is a multiple of 3 (for the codons).

Let me know if that clarifies things, or if you have other concerns. The breaches are evaluated and solved relatively independently, see readme for a gif demonstration: https://github.com/Edinburgh-Genome-Foundry/DnaChisel?tab=readme-ov-file#how-it-works

Of course you can specify additional constraints with AvoidChanges for confirmation that your N and C terminals pass, or objectives if you want set something globally on the whole sequence, for example GC content.