De novo assembly, gene annotation and marker development using illumina paired-end transcriptome sequences in nutmeg ( Houtt).

Nutmeg () is a high-value spice crop widely utilized in culinary, medicinal, and industrial sectors. Despite its economic and cultural significance, genetic research on nutmeg is severely limited, hindering efforts to improve its cultivation and breeding programs. The lack of molecular markers and genetic tools has further constrained the development of superior trait varieties. Molecular markers are essential for genetic mapping, diversity studies, and marker-assisted selection, yet their application in nutmeg remains unexplored. This study utilized Illumina high-throughput paired-end transcriptome sequencing to explore the genetic framework of nutmeg, focusing on de novo assembly, gene annotation, and molecular marker development. RNA Sequencing (RNA-seq) was performed on leaf tissues from male and female trees to analyze differentially expressed genes. A total of 141,192 unigenes were assembled with an average length of 1117 bp, achieving functional annotation in public databases like NR, Swiss-Prot, KEGG, and InterPro. De novo transcriptome assembly and functional annotation revealed ten candidate genes involved in sex determination. These candidate genes are associated with hormonal pathways, transcription factors, and reproductive development. Several primers/markers have been designed and validated using PCR. This research lays a foundation for genetic studies, marker-assisted breeding, and improved cultivation practices for nutmeg.