Analysis of codon bias in WRKY transcription factors related to seawater stress in Hemerocallis fulva

During the process of plant evolution, genes of different species exhibit specific codon usage biases. The WRKY gene family plays an important role in plant stress resistance. Hemerocallis fulva is an excellent horticultural plant with strong stress resistance. To further explore the codon usage characteristics of Hemerocallis fulva WRKY (HfWRKY) genes, the CodonW and EMBOSS online programs were employed to analyze 21 WRKY transcription factors related to seawater stress induction. Additionally, Excel and Rstudio software were utilized to generate neutrality analysis charts, ENc-plot charts, and PR2-plot charts, aiming to reveal the codon usage patterns of HfWRKY genes. The results showed that the average GC content of HfWRKY genes was 48.31%, while the average GC3 content was 51.72%, indicating that HfWRKY genes tend to use G/C-ending codons with a relatively weak codon usage bias. Relative Synonymous Codon Usage (RSCU) analysis revealed that 28 codons had RSCU values greater than 1, with 13 ending in A or T and 15 ending in G or C. Furthermore, the effective number of codons ranged from 46.54 to 59.64, and the codon adaptation index ranged from 0.158 to 0.274. Through neutrality analysis, ENc-plot, and PR2-plot charts, it was found that the codon usage patterns of HfWRKY genes were influenced by factors such as mutation and natural selection. Finally, 10 optimal codons were identified in the HfWRKY gene family, demonstrating their association with resistance. The results of this study provide a theoretical basis for further exploring the role of the HfWRKY gene family in seawater stress resistance and molecular breeding in Hemerocallis fulva.