Background Camptotheca acuminata is a Nyssaceae place, often called the “happy tree”, which is indigenous in Southern China. (CaPSTR) were cloned 1202916-90-2 IC50 and analyzed. The expression level of the three genes was also detected using qRT-PCR in C. acuminata. With respect to the branch pathway of CPT synthesis, six cytochrome P450s transcripts were selected as candidate transcripts by detection of transcript 1202916-90-2 IC50 expression in different tissues using qRT-PCR. In addition, one glucosidase gene was recognized that might participate in CPT biosynthesis. For CPT transport, three of 21 transcripts for multidrug resistance Nrp1 protein (MDR) transporters were also screened from your dataset by their annotation result and gene expression analysis. Conclusion This study produced a large amount of transcriptome data from C. acuminata by 454 pyrosequencing. According to EST annotation, catalytic features prediction, and expression analysis, novel putative transcripts involved in CPT biosynthesis and transport were discovered in C. acuminata. This study will facilitate further identification of important enzymes and transporter genes in C. acuminata. Background Camptothecin (CPT) was first extracted from your 1202916-90-2 IC50 stems of Camptotheca acuminata in 1966 and subsequently from Nothapodytes foetida, Ophiorrhiza pumila, and Ophiorrhiza japonica [1]. CPT exhibits clinical anti-tumor activity by inhibiting DNA topoisomerase I, an enzyme involved in DNA recombination, repair, replication, and transcription [2]. CPT also inhibits the retroviruses, such as the human immunodeficiency computer virus [3]. Despite its significant clinical use, the main source of CPT is still from its extraction from C. acuminata. However, the quantity is quite limited and cannot meet worldwide demand. Studies around the molecular mechanism of CPT biosynthesis have long been hindered by the lack of transcriptome and genome information for C. acuminata and other CPT-producing plants. Therefore, it is necessary to obtain transcriptome data and screen candidate transcripts involved in CPT biosynthesis to further understand the CPT biosynthetic pathway. CPT is usually synthesized through a altered terpenoid indole alkaloid (TIA) pathway. The upstream biosynthesis pathways for all the TIA products are comparable among alkaloid-producing plants, and involve a strictosidine backbone (Physique ?(Figure1A).1A). Over recent decades, several enzymes in the process of strictosidine biosynthesis in C. acuminata have been isolated and functionally recognized. Among them are tryptophan synthase (TSB) [4] and tryptophan decarboxylase (TDC) [5], which are involved in the synthesis of the indole precursor tryptamine, 3-hydroxy-3-methylglutaryl-CoA synthase (HMGR) [6], 1-deoxy-D-xylulose-5-phosphate reductoisomeras (DXR) [7], and 10-hydroxy geraniol oxidoreductase (10HGO) [8] are involved in secologanin synthesis. Physique 1 Biosynthetic pathway of CPT from DMAPP to strictosidine and from strictosidine to CPT in C. acuminata. (A) The upstream pathway for the synthesis of backbone strictosidine. (B) The proposed branch pathway of CPT biosynthesis (actions after strictosidine … G10H and SCS, belonging to the CYP76B6 and CYP72A1subfamilies of cytochrome P450 family respectively, were recognized in monoterpenoid biosynthesis from Catharanthus roseus [9,10]. The synthesis of strictosidine is usually finally catalyzed by STR, a 1202916-90-2 IC50 committed enzyme for the CPT backbone biosynthesis, which was isolated and recognized in Rauvolfia serpentine, C. roseus, the CPT-producing herb O. japonica, and O. pumila, in previous studies. However, the genes encoding CaG10H, CaSCS and CaSTR, have not been yet cloned and characterized in C. acuminata. The actions following strictosidine formation (branch pathway) are not very clear and only a proposed biosynthetic pathway based on relative compounds extracted from CPT-producing plants has been reported [11] (Physique ?(Figure1B).1B). In the proposed pathway, a series of oxidation and hydroxylation reactions are involved in some steps of the pathway which are probably catalyzed by monooxygenases and hydroxylase, belonging to the superfamily of cytochrome P450s [12,13]. In the mean time, the.