| Wood identification is meaningful for wood trading, wood processing, archaeologyof woody relic, wood inspecting and quarantining, disputes and some cases referringwood or wood products, which have been taking into account by certain offices andpeople. Authoritative wood identification depend on specialized department and staff.Generally, analysing its anatomical characteristics is the traditional means for woodidentification. While, different identifiers may make different conclusions for the samewood sample, especially for some wood samples which get similar characters. Sotraditional wood identification demands identifier high level of professional quality andabundant data of various wood anatomical characteristcs. However, traditional way ofwood identification may encounter the challenge to species level.DNA barcode would be the unique identifier for species by selecting certainsequence of its genetic material. Other than traditional means, DNA barcoding techniquecan reveal the differences of the species in genetic level, which is accurate. DNAbarcoding gets advantages: convenient and timesaving procedure for identication, adaptto test large number of samples, high stability and repeatability, standardized testprocedure, and sharing international database for analysis.In this study, DNA barcoding was used in wood identification. Two precious woodspecies: Dalbergia odorifera T. Chen and Dalbergia rimosa Roxb which origined fromHainan, Guangxi and Guangzhou were selected as material. Three methods i.e. modifiedCTAB, modified QIAGEN kit, modified PTB method were compared to isolate DNAfrom wood. And4candidate barcodes (matK, rbcL, trnH-psbA, ITS2) which wererecommonded by Barcode of Life Data Systems were assessed by the success rate ofPCR amplication and sequencing of each segment. Further evaluation was made byanalyzing each candidate barcode sequences information. This research would provideinformation for wood identification via DNA barcoding, and for constracting barcodingdatabase of Chinese rosewood.The results showed that, DNA from heartwood can be extracted with modified PTBmethod effectively. And the DNA concentration ranged from0.34~3.52ng/μL. While,DNA from sapwood can be isolated effectively by modified CTAB method and the DNAconcentration reached185.45ng/μL. It is confrimed that DNAextraction from heartwoodis much more difficult than from sapwood. The4candidate barcodes were analysed.Success rate of PCR amplication of matK was only25%. Success rate of PCR amplication and sequencing of rbcL were100%and75%respectively. Success rate ofPCR amplication and sequencing of trnH-psbA were91.67%and100%. Success rate ofPCR amplication and sequencing of ITS2were66.67%and87.5%. The length of rbcLwas703bp,(G+C)%was42.4%,(A+T)%was57.6%, however, no variation wereobserved both in Dalbergia odorifera T. Chen and in Dalbergia rimosa Roxb. The lengthof trnH-psbA ranged from318-322bp,(A+T)%was about52%,(G+C)%was about48%,Intraspecific genetic distance of Dalbergia odorifera T. Chen and Dalbergia rimosaRoxb was0.0059and0.0076respectively. Interspecific genetic distance between the twospceies was0.0066. So there was no specific DNA barcoding gap existing. ITS2was482-483bp,(G+C)%was about61%，(A+T)%was about38%. The genetic distancebetween Dalbergia rimosa Roxb was0.007. The genetic distance between Dalbergiaodorifera T. Chen and Dalbergia rimosa Roxb was0.0119. The secondary structure ofITS2of the two wood species were obtained and compared. The structure of the twospecies were vary similar, got the same reverse angle. The main difference between thetwo species was the Helix4. Phylogenetic tree of ITS2was constructed. Dalbergiaodorifera T. Chen and Dalbergia rimosa Roxb were divergence to separated branchs.The secondary structure and phylogenetic tree all revealed that the two species ofDalbergia odorifera T. Chen and Dalbergia rimosa Roxb were high related but separated.What is more, ITS2was commended as candidate DNAbarcode in this research. |
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