Mutanalyst

Mutanalyst analyzes mutational spectra of libraries generated by error-prone PCR (epPCR) to quantify nucleobase substitution proportions in sequenced colonies and estimate the mean number of mutations per sequence for directed evolution experiments.

Key Features:

• Automated Mutational Spectrum Calculation: Automates calculation of mutation spectra by determining proportions of specific nucleobase changes within sequenced colonies from an epPCR library.
• Error Estimation through DNA Complementarity: Estimates errors in mutational spectrum calculations by leveraging DNA complementarity between strands to refine mutation counts.
• Robust Statistical Analysis with Poisson Distribution: Fits mutation count data to a Poisson distribution to determine the mean number of mutations per sequence and improve robustness for small sample sizes.
• Guidance for Experimental Adjustments: Provides quantitative metrics to assess whether epPCR conditions yield desired mutation diversity and to inform adjustments to experimental parameters.

Scientific Applications:

• Directed Evolution Studies: Characterizes library diversity and mutation distributions to support selection and screening strategies in directed evolution experiments.
• Protein Engineering Optimization: Quantifies mutation types and rates to guide library design and increase the likelihood of identifying beneficial substitutions in protein engineering.
• Library Quality Assessment and Experimental Design: Assesses adequacy of epPCR-generated libraries and aids design decisions to improve coverage of sequence space.

Methodology:

Calculates proportions of nucleobase substitutions from sequenced colonies, estimates errors via DNA complementarity between strands, and fits mutation counts to a Poisson distribution to infer the mean mutations per sequence.