Developing SSR Individual Identification System and Tracking the Geographical Origin of Timber for the Endangered Species Chamaecyparis Formosensis

Chamaecyparis formosensis is an endemic species of Taiwan, threatened from intensive use and illegal felling. An individual identification system for C. formosensis is required to provide scientific evidence for court use and deter illegal felling. In this study, 36 polymorphic simple sequence repeat (SSR) markers were developed. By applying up to 28 non-linkage of the developed markers, it is calculated that the cumulative random probability of identity (CPI) is as low as 1.652×10-12, the combined power of discrimination (CPD) is as high as 0.999999999998348, and the identifiable population size is up to 60 million, which is greater than the known C. formosensis population size in Taiwan. Biogeographical analysis data show that C. formosensis from four geographic areas belong to the same genetic population which can be further divided into three clusters: SY (Eastern Taiwan), HV and GW (Northwestern Taiwan), and MM (Southwestern Taiwan). The developed system was applied to assess the provenance of samples with 88.44% of accuracy rate, and therefore can serve as a pre-screening tool to reduce the range required for comparison. The system developed in this study is a potential crime-fighting tool against illegal felling.

Genome skimming and exploration of DNA barcodes for Taiwan endemic cypresses

Cypresses are characterized by their longevity and valuable timber. In Taiwan, two endemic cypress species, Chamaecyparis formosensis and C. obtusa var. formosana, are threatened by prevalent illegal logging. A DNA barcode system is urgently needed for reforestation and conservation of these two cypresses. In this study, both plastomes and 35S rDNAs from 16, 10, and 6 individuals of C. formosensis, C. obtusa var. formosana, and C. obtusa var. obtusa were sequenced, respectively. We show that the loss of plastid trnT-GGU readily distinguishes C. formosensis from its congeneric species. We demonstrate that entire sequences of plastomes or 35S rDNAs are capable of correctly identifying cypress species and varieties, suggesting that they are effective super-barcodes. We also discover three short hypervariable loci (i.e., 3′ETS, ITS1, and trnH-psbA) that are promising barcodes for identifying cypress species and varieties. Moreover, nine species-specific indels of > 100 bp were detected in the cypress plastomes. These indels, together with the three aforementioned short barcodes, constitute an alternative and powerful barcode system crucial for identifying specimens that are fragmentary or contain degraded/poor DNA. Our sequenced data and barcode systems not only enrich the genetic reference for cypresses, but also contribute to future reforestation, conservation, and forensic investigations.

Composition, in vitro Anti-mildew Fungal Activities of the Heartwood Essential Oil of Chamaecyparis formosensis from Taiwan

In this study, anti-mildew fungal activities of the heartwood essential oil, and its constituents from Chamaecyparis formosensis were evaluated in vitro against 7 mildew fungi. The main compounds responsible for the anti-mildew fungal activities were isolated and identified. The heartwood essential oil of C. formosensis was isolated using hydrodistillation in a Clevenger-type apparatus, and characterized by GC-FID and GC-MS. The heartwood oil consisted primarily of myrtenol (18.4%), cis-myrtanol (14.0%), α-muurolol (13.8%), α-cadinol (12.7%), and chamaecynone (9.8%). The heartwood oil was shown to have excellent anti-mildew fungal activities. Further fractionation of the heartwood oil produced α-cadinol, chamaecynone, α-muurolol, τ-cadinol, and τ-muurolol. The 5 compounds exhibited very strong anti-mildew fungal activities. For the anti-mildew fungal activities of the heartwood oil, the active source compounds were determined to be α-cadinol, chamaecynone, α-muurolol, τ-cadinol, and τ-muurolol.