## 3 Discussion  
Fungi generate a diverse range of bioactive secondary metabolites, which could be associated with their function as decomposers in ecosystems, as well as their role as plant mutualists and nutrient-rich food sources for numerous insects [4; 31]. Thus, they have developed the capacity to generate large quantities of compounds that possess insecticidal, alluring, and even carnivorous properties during their long struggle against insects [5; 32]. For example, the indole-diterpenoid compound peintrem G, isolated from _P. crustosum_, shows convulsive and insecticidal activities against _Oncopelus fasciatus_ and _Ceratitis capitata_[33]. Pyripyropene A, extracted from _P. coprobium_ and _P. reticulporum_, inhibits acetylcholinesterase and possesses significant insecticidal efficacy against aphids. It serves as a prototype for the development of novel insecticides, including Afidopyropen [34]. The compound 2-furoic acid has nematicidal properties. It effectively poisoned nematodes upon infection and significantly enhances the nematode-capturing ability of _Dactylellina haptotyla_[35]. Despite the discovery of a considerable number of insecticidal metabolites in fungi (e.g., more than 150 compounds in _Penicillium_ spp. [5]), only a minority have progressed to the stage of development into insecticides. Additional in-depth research on insecticidal activity, spectrum, environmental safety, and development of insecticide products is significantly impeded by the capacity to manufacture large quantities of naturally or artificially synthesized pesticides. Understanding the biosynthetic gene clusters (BGCs) of secondary metabolites produced by fungi has rendered the transcriptional regulation of such BGCs a crucial factor in elucidating the efficacy of product biosynthesis. A substantial variation in the yield of Okaramine B was observed because of fermentation conducted under distinct culture conditions. Using comparative transcriptomics, this study sought to investigate the transcriptional differences of _oka_ cluster and regulatory-related genes. Approximately 50% of fungal BGCs contain a transcription factor unique to the cluster. This cluster-specific transcription factor is often a C6-zinc cluster protein that can identify palindromic patterns in the promoters of genes inside the cluster [4; 36]. In this study, the gene cluster _oka_ of Okaramine B did not contain or flank regulatory-related genes [21]. In addition to cluster-specific transcription factors, secondary metabolite regulation also depends on global regulation. The Velvet complex, consisting of LaeA, VeA, and VelB, is the most influential transcriptional complex known to affect the global regulation of secondary metabolites in fungi [37]. In the transcriptome analysis conducted in this study, gene 4.839, encoding the protein with 66.74% aa identity with LaeA in _P. rubens_ (accession no. B6H9U8.1), exhibited transcriptional up-regulation starting from the fourth day. However, no significant transcriptional alterations were observed in genes 4.494 or 1.65. These two genes encode proteins that share 75.0% and 72.1% aa identity, respectively, with a sexual development activator VeA in _P. cittrinum_ (accession no. BAL61195.1) and VelB in _P. rubens_ (accession no. B6 HU70.1). Similar cases that exhibited non-differential expression included the following: gene 1.2132, the protein of which shares 52.9% aa identity with the global negative regulator McrA of BGCs in _Penicillium_ and _Apergillus_ species [38]; gene 1.1524, the protein of which shares 77.1% aa identity with the Zn(II)2-Cys6-type regulator KpeA in _A. oryzae_ (accession no. Q2 UJ4.1) [39]; gene 5.211, the protein of which shares 76.9% aa identity with Myb domain-containing transcription factor in _Hapsidospora chrysogene_ (accession no. AIZ05820.1) [40]; and so on. However, it is interesting to note that this study identified, in addition to the homologs of the regulator BrIA (gene 4.1080), some regulatory genes that potentially influence the biosynthesis of Okaramine B.