Genome-Wide Identification of the PAL Gene Family in Camellia nitidissima and Functional Characterization of CnPAL1 Gene by In Vitro Expression

<b>Background</b>: <i>PAL</i> genes are crucial for plant growth and stress response, yet studies on the <i>PAL</i> gene family in <i>Camellia nitidissima</i> are sparse. <b>Methods</b>: The <i>PAL</i> gene family was screened from the entire genome of <i>C. nitidissima</i>, and their physicochemical properties, chromosomal locations, intraspecific and interspecific collinearity, conserved motifs, phylogenetic trees, cis-acting elements, and gene structures were analyzed. The expression patterns of the <i>CnPAL</i> genes were compared across different tissues, and the highly expressed <i>CnPAL1</i> gene was expressed in prokaryotes, and its enzyme activity was validated using UPLC-MS technology. <b>Results</b>: The results revealed that six <i>CnPALs</i> were identified in the <i>C. nitidissima</i> genome, distributed unevenly across six chromosomes. The CnPAL proteins shared similar physicochemical properties, with highly conserved motifs and gene structures. Promoter analysis showed multiple cis-acting elements in the <i>CnPALs</i> genes. Intra-species collinearity analysis revealed that all <i>CnPALs</i> were collinear with multiple <i>PAL</i> genes in <i>C. nitidissima</i>, while inter-species collinearity analysis indicated that <i>CnPALs</i> were collinear with the <i>PAL</i> genes in <i>Camellia oleifera</i> and <i>Camellia sinensis</i>. Furthermore, the transcriptomic data of <i>C. nitidissima</i> demonstrated tissue-specific expression of the <i>CnPALs</i>, although qRT-PCR validation showed some discrepancies with the sequencing result. The qRT-PCR revealed varied expression patterns among the six <i>CnPALs</i>, with the <i>CnPAL1</i> gene showing relatively higher expression levels. Subsequently, cloning, prokaryotic expression, and enzyme activity analysis confirmed the effective catalytic activity of the CnPAL1 protein. <b>Conclusions</b>: This study lays the foundation for understanding the functions of <i>CnPAL</i> genes and offers insights for genetic improvement of <i>C. nitidissima</i>.