Siregar Iskandar Z, Ramdhani Muhammad Jauhari, Karlinasari Lina, Adzkia Ulfa, Arifin M Zainul, Dwiyanti Fifi Gus
Department of Silviculture, Faculty of Forestry and Environment, IPB University, IPB Dramaga Campus, Dramaga, Bogor, Indonesia.
Department of Silviculture, Faculty of Forestry and Environment, IPB University, IPB Dramaga Campus, Dramaga, Bogor, Indonesia.
Sci Justice. 2021 Sep;61(5):573-578. doi: 10.1016/j.scijus.2021.07.002. Epub 2021 Jul 8.
The successful isolation of DNA (Deoxyribonucleic Acid) is essential for the investigation process of forestry molecular genetics. Samples used are usually retrieved either from soft or juvenile plant organs because of their excellent DNA source. However, in certain cases, aforesaid samples are hard to obtain, as for forensic purposes. Alternatively, woods possess potential as alternative source of DNA whose extraction method has been developed with varying degrees of success. However, to date, effectiveness on tropical wood grown in Indonesia has not been widely reported. Therefore, objective of this study was to compare the results of DNA isolation of various dried and fresh wood samples by using two isolation methods: Cetyl Trimethyl Ammonium Bromide (CTAB) and Qiagen DNeasy Plant Mini Kit (QDPMK). Extraction results were visualized in agarose gels and quantified using Nanophotometer NP80 Implen which were then amplified using two universal primers: ITS and rbcL for detecting DNA signals. Extraction results from dried wood indicated no visualization in the gel, while fresh wood samples showed thick smeared bands on both extraction methods. Quantity test results denoted higher concentration in CTAB-extracted samples compared to samples extracted using QDPMK, in both types of samples, even though both resulted in optical density ratios outside the range of purity (λ260/280: 1,8-2,0 and λ260/230: 2,0, respectively). Success rates of ITS and rbcL primary amplification in dried wood samples were quite low yet outputs from the two methods did not differ significantly. Meanwhile, outcome of ITS and rbcL amplification on fresh wood samples had a fairly high success.
成功分离DNA（脱氧核糖核酸）对于林业分子遗传学的研究过程至关重要。由于其作为优质DNA来源，所使用的样本通常取自软质或幼年植物器官。然而，在某些情况下，出于法医目的，上述样本难以获取。另外，木材具有作为DNA替代来源的潜力，其提取方法已得到不同程度的开发。然而，迄今为止，关于印度尼西亚种植的热带木材的提取效果尚未有广泛报道。因此，本研究的目的是通过使用两种分离方法：十六烷基三甲基溴化铵（CTAB）和Qiagen DNeasy植物微量提取试剂盒（QDPMK），比较各种干燥和新鲜木材样本的DNA分离结果。提取结果在琼脂糖凝胶中可视化，并使用Nanophotometer NP80 Implen进行定量，然后使用两种通用引物：ITS和rbcL进行扩增以检测DNA信号。干燥木材的提取结果在凝胶中未显示可视化，而新鲜木材样本在两种提取方法中均显示出浓重的拖带。定量测试结果表明，在两种类型的样本中，CTAB提取的样本浓度均高于使用QDPMK提取的样本，尽管两者的光密度比均超出纯度范围（分别为λ260/280：1.8 - 2.0和λ260/230：2.0）。干燥木材样本中ITS和rbcL初次扩增的成功率相当低，但两种方法的输出结果差异不显著。同时，新鲜木材样本上ITS和rbcL扩增的结果成功率相当高。
