Objective The plant NHX (Na⁺/H⁺ exchanger) gene family encodes Na⁺/H⁺ antiporter proteins, which are widely involved in the response to salt stress and the reconstruction of ion balance in plants, playing a crucial role in plant salt tolerance. This study aims to identify and analyze the NHX gene family in Tetragonia tetragonoides to explore its role in the salt tolerance of this species. T. tetragonoides is a wild halophytic vegetable with strong salt tolerance, commonly found on tropical and subtropical beaches.

Method Based on the whole-genome sequencing data of T. tetragonoides, 13 TtNHX genes were identified using bioinformatics approaches. A comprehensive analysis was conducted on their chromosomal localization, gene structure, promoter cis-acting elements, protein characteristics, and evolutionary relationships. The expression profiles of TtNHX genes were preliminarily tested by RNA sequencing (RNA-seq) and RT-qPCR assays.

Result A total of 13 members of the TtNHX gene family were identified, distributed across 11 chromosomes. The coding regions of these 13 TtNHX proteins range in length from 521 to 1, 146 amino acids, with molecular weights ranging from 58.14 to 127.06 kD and isoelectric points ranging from 5.17 to 8.79. Phylogenetic tree results indicate that TtNHX protein members in T. tetragonoides exhibit high conservation, clustering with NHX proteins from Arabidopsis, rice, tomato, and Vicia faba into three major subfamilies (plasma membrane-type, endoplasmic reticulum-type, and vacuolar-type), with notable duplication and expansion among their members. RNA-seq and quantitative RT-qPCR results indicate that members of the TtNHX gene family are widely and differentially expressed across different organs of T. tetragonoides. Among them, TtNHX2/TtNHX13 and TtNHX4/TtNHX8 showed higher expression in roots and stems, while TtNHX1/TtNHX11 and TtNHX7/TtNHX12 exhibited lower expression across organs. These findings suggest potential roles in regulating T. tetragonoides development and responses to various abiotic stresses.

Conclusion 13 TtNHX gene family members were identified. This research provided the theoretical foundation for identification of key functional salt-tolerance genes, as well as identified the excellent candidate resistance genes for molecular breeding of salt resistance in crops, including T. tetragonoides and other vegetables.