The high-throughput sequencing results will also reveal other pathways that are related to wax synthesis and will identify a larger Vemurafenib number of polymorphism molecular markers, which are scarce in the Welsh onion. Based on the RNA-seq data, the closely related gene expression patterns were investigated to illustrate the function of these genes in the wax synthesis pathway. This research will provide additional evidence of waxy gene expression in wax synthesis and can be used to develop methods for mapping waxy genes and other genes in the Welsh onion. To obtain high-quality clean read data for de novo assembly, the raw reads were filtered by removing the adaptor sequences, duplication sequences, reads with an “N” rate greater than 10%, and low-quality reads with more than 50% of the bases with a Q-value #5. The clean reads were assembled into contigs using the Trinity method, which efficiently reconstructs full-length transcripts across a broad range of expression levels and sequencing depths. Contigs were created by combining reads that had a certain length of overlap. The reads were then mapped back to the contigs; with paired-end reads itis able to detect contigs from the same transcript as well as the distances between these contigs. The contigs were connected using the Trinity software to get sequences that could not be extended at either end. Such sequences were defined as unigenes, and the unigenes were combined to produce the final assembly used for annotation. We quantified the transcript levels in reads per kilobase of the exon model per million mapped reads. The RPKM measure of read density reflects the molar concentration of a transcript in the starting sample by normalizing the RNA length and the total read number in the measurement. By using Bowtie, each sequencing read sample was compared with the UniGene database and, using RPKM, reflected the expression abundance of the unigenes. With the improvement of read length by paired-end sequencing, relatively short reads can be effectively assembled and have been successfully used to study plants without a genomic sequence. The study of Kolattu-kudy provides insights into the basic information necessary to analyze and identify the products that are synthesized by the waxy gene. Due to the paucity of studies on wax-related genes in the Welsh onion, the important waxy metabolism genes in the Welsh onion are not known. This study used the Illumina HiSeq 2000 platform for RNA-Seq to profile the Welsh onion transcriptome. In addition, the functions of the unigenes were classified by the COG and GO annotations and the metabolic pathways. A total of 798 genes, representing 1.86% total putative unigenes, were differentially expressed between the waxy Welsh onion and non-waxy mutant Welsh onion varieties. Through SwissProt annotation, four important waxy synthetic unigenes of Welsh onion were found, and the results showed that the waxy synthesis protein might be controlled by Protein ECERIFERUM 3, Long-chain acyl-CoA synthetase 2 waxy gene synthesis in lipid transport and metabolism, and the ABC transporter G family member 12 in the defense mechanism.