Applying nephelometry for analyzing liquid yeast cultures

Saccharomyces cerevisiae is a widely used model organism for the molecular analysis of genes and proteins. Several methods have been developed to study protein function and activity through heterologous gene expression, including yeast two-hybrid and yeast complementation. Traditionally, these yeast-based assays were performed on solid agar plates. While this approach provides an easy visual readout, it is difficult to quantify the results accurately. To overcome this limitation, liquid-based methods were introduced. Most of these methods rely on the use of spectrophotometry to measure reduction in light transmission as a result of light scattering and monitor culture growth. In this study, we propose nephelometry as an additional method for performing and analyzing liquid-culture yeast complementation assays. More specifically, we compare the suitability of using nephelometry for the functional analysis of two homologous proteins using yeast complementation: The amino acid transporter homologues Arabidopsis thaliana LYSINE HISTIDINE TRANSPORTER 1 (AtLHT1) and Populus tremula L. x tremuloides Michx LYSINE HISTIDINE TRANSPORTER 1.2 (PtrLHT1.2). In previous reports, no differences in microbial growth were detected, irrespective of which homolog was used to rescue an amino acid-deficient yeast mutant strain. By using nephelometry to record yeast growth, we demonstrated that it is a robust and reproducible method. When comparing to spectrophotometric measurements of yeast cultures, it proved to be a suitable alternative. The novel approach even revealed previously undetected differences in culture growth of both homologues, highlighting nephelometry's potential to improve sensitivity in yeast-based functional assays. We present the use of nephelometry as an equal method to yeast complementation traditionally executed on solid agar medium or in liquid culture with spectrophotometric analysis.