Engineering proteins with optimized function requires exploring large and diverse sequence spaces, often through the construction and screening of thousands of long gene variants. However, assembling genes exceeding ~1 kb—particularly when variants are distributed across the sequence and generated in pooled formats—remains technically challenging. Conventional assembly methods can introduce errors, uneven representation, and chimeric constructs that limit library quality and slow experimental progress.

In this white paper, we describe an approach for generating accurate, uniform gene libraries up to 1.8 kb in length using Twist Bioscience’s Gene Pools technology. By enabling high-fidelity assembly of complex variant libraries while minimizing chimera formation, this method supports more efficient and scalable protein engineering workflows.

Covered in this White Paper: