| Summary: | 碩士 === 國立中山大學 === 海洋生物研究所 === 90 === Abstract
This study is to elucidate the possibility of DNA fragment identification method in the forensic detection of marine microalgae (Chlorella sp. and Nannochloropsis oculata) from human (Homo sapiens) body by 3% agarose gel electrophoresis of amplification fragments via PCR-amplified ribosomal gene small subunit (SSU) rDNA molecules as primers, which are specific nucleotide segments on conserved regions (18S rDNA region (NS1-NS2 primers))on SSU rDNA and on variable regions (internal transcribed spacer (ITS) regions (primer 2/primer 5 for ITS1 region and primer 3/primer 4 for ITS2 region)). NS1-NS2-amplified PCR fragments are 550 bp for C. sp., 550 bp for N. oculata and 600 bp for human. ITS1-amplified PCR fragments are 1, 2 and 4 bands for C. sp. (350 bp), N. oculata (400 and 450 bp) and human (300, 450, 500 and 580 bp), respectively, while ITS2-amplified PCR fragments are 1, 2 and 1 bands for C. sp. (430 bp), N. oculata (430 and 600 bp) and human (400 bp), respectively. By using human-specific primers (AmpFlSTR® Profilerä PCR Amplification Kit), only human can be identified in the sample containing C. sp., N. oculata and human DNA, whereas C. sp. and N. oculata can not be detected, indicating the prevention of algal interference in human-specific primer-PCR procedures via AmpFlSTR® Profilerä PCR Amplification Kit. Detection limits of C. sp. and N. oculata DNA were 50 and 10 pg, respectively. The results of present investigation show that algae can be distinguished from human by NS1-NS2-amplified PCR fragments but not between C. sp. and N. oculata, while C. sp., N. oculata and human can be distinguished by ITS1- or ITS2-amplified PCR fragments. Evidently, the specificity of DNA segments in marine microalgal and human DNA provides the base for investigation of cause of death in drowning case in the marine environment.
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