Sodium Transport Function Yeast Assay

Service Introduction

Specifically designed for the study of yeast sodium metabolism and transport protein function, based on the principle of heterologous complementation experiments in yeast. It includes sodium-deficient yeast strains, expression vectors, and supporting reagents. Sodium-deficient strains, due to the absence of endogenous transport proteins, show inhibited growth in high-sodium environments. After transforming the strain with the target transport protein gene, if the protein is active, it can complement the defective phenotype, directly reflecting transport capability and excluding interference from other cations. This system is suitable for verifying the function of transport proteins, screening sodium metabolism genes, and studying regulatory mechanisms. The AXT3K Saccharomyces cerevisiae strain is a Na+/sodium-deficient strain, lacking the endogenous plasma membrane Na+ antiporter NHX1 as well as Na+ efflux proteins NHA1 and ENA1-4, resulting in the loss of Na+ transport capability. AXT3K is widely used in Na+ transport protein identification experiments and can also be used for identifying Na+ channels or sodium-potassium pumps. This strain is particularly sensitive to Na+, showing significantly inhibited growth on media containing 50 mM NaCl. pYES2 is a high-efficiency expression vector for Saccharomyces cerevisiae proteins, with Ura3 as the selection marker. The GAL1 promoter requires galactose to induce protein expression. By constructing the Na+ transport protein gene X into the pYES2 vector (pYES2-X) and transforming it into AXT3K competent cells, protein X will mediate yeast to expel excess sodium ions into the medium, allowing AXT3K yeast to grow normally on high-sodium AP media.

Service Advantages

▶Can detect whether a protein has sodium ion uptake and transport activity, simple and efficient.

▶Our company has all the reagents, strains, and vectors needed for detecting sodium ion transport activity.

Service Process

Article Case

Reconstitution in yeast of the Arabidopsis SOS signaling pathway for Na homeostasis

academic journal：PNAS

SOS2 and SOS3 increase the NaCl tolerance of yeast expressing the SOS1 ion transporter. AXT3K cells transformed with an empty vector (control) or expressing the indicated combination of Arabidopsis genes were grown overnight in liquid AP medium with 1 mM KCl. Five microliters of serial decimal dilutions were spotted onto plates of the same medium or supplemented with 70 mM NaCl. Plates were incubated at 28°C and photographed after 4 days. Plasmids used for expression of the SOS proteins were: pSOS1–1 for SOS1; pFL2T for SOS2; pFL3T for SOS3; pFL32T for SOS2 and SOS3; and pFL2△T for SOS2T D△308.

Applicable Fields

▶Functional identification of sodium transport proteins: Determine the in vivo transport function of candidate sodium transporter genes from plants, microorganisms, and animals, and clarify whether the target gene possesses the core activity to mediate transmembrane sodium ion uptake. This is a key technical approach for gene function validation.

▶Plant stress physiology research: Analyze the role of sodium transport proteins in maintaining intracellular sodium homeostasis in crops under stresses such as sodium deficiency and heavy metal exposure; screen for crops with low-sodium tolerance or high sodium utilization efficiency, providing targets for nutrient-efficient breeding.

▶Microbial sodium metabolism mechanism analysis: Study the sodium transport systems in bacteria, fungi, and other microorganisms, clarifying the regulatory mechanisms of sodium transport proteins in microbial adaptation to sodium-deficient environments and in maintaining enzyme activities (e.g., sodium superoxide dismutase).

▶Functional screening of sodium transporter mutants/variants: Validate the functions of different mutants and allelic variants of sodium transport proteins, assess the impact of amino acid site mutations on transport activity, and support studies on the relationship between protein structure and function.