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干态 DNA 室温储存及运输方案

Protocols for dry DNA storage and shipment at room temperature.

机构信息

Biodiversity Institute of Ontario, University of Guelph, Guelph, ON, N1G 2W1, Canada.

出版信息

Mol Ecol Resour. 2013 Sep;13(5):890-8. doi: 10.1111/1755-0998.12134. Epub 2013 Jun 24.

Abstract

The globalization of DNA barcoding will require core analytical facilities to develop cost-effective, efficient protocols for the shipment and archival storage of DNA extracts and PCR products. We evaluated three dry-state DNA stabilization systems: commercial Biomatrica(®) DNAstable(®) plates, home-made trehalose and polyvinyl alcohol (PVA) plates on 96-well panels of insect DNA stored at 56 °C and at room temperature. Controls included unprotected samples that were stored dry at room temperature and at 56 °C, and diluted samples held at 4 °C and at -20 °C. PCR and selective sequencing were performed over a 4-year interval to test the condition of DNA extracts. Biomatrica(®) provided better protection of DNA at 56 °C and at room temperature than trehalose and PVA, especially for diluted samples. PVA was the second best protectant after Biomatrica(®) at room temperature, whereas trehalose was the second best protectant at 56 °C. In spite of lower PCR success, the DNA stored at -20 °C yielded longer sequence reads and stronger signal, indicating that temperature is a crucial factor for DNA quality which has to be considered especially for long-term storage. Although it is premature to advocate a transition to DNA storage at room temperature, dry storage provides an additional layer of security for frozen samples, protecting them from degradation in the event of freezer failure. All three forms of DNA preservation enable shipment of dry DNA and PCR products between barcoding facilities.

摘要

DNA 条码的全球化将需要核心分析设施,以开发具有成本效益、高效率的协议,用于运输和存档 DNA 提取物和 PCR 产物。我们评估了三种干态 DNA 稳定系统:商业的 Biomatrica(®) DNAstable(®) 板、自制的海藻糖和聚乙烯醇 (PVA) 板,用于存储在 56°C 和室温下的昆虫 DNA 的 96 孔板。对照包括在室温下和 56°C 下干燥保存的未保护样品,以及在 4°C 和 -20°C 下保存的稀释样品。PCR 和选择性测序在 4 年的时间间隔内进行,以测试 DNA 提取物的状况。Biomatrica(®) 在 56°C 和室温下比海藻糖和 PVA 更能保护 DNA,尤其是对稀释的样品。PVA 在室温下是 Biomatrica(®) 之后的第二好的保护剂,而海藻糖在 56°C 下是第二好的保护剂。尽管 PCR 成功率较低,但储存在 -20°C 的 DNA 产生了更长的序列读长和更强的信号,表明温度是 DNA 质量的关键因素,尤其是对于长期储存,必须考虑到这一点。虽然现在提倡在室温下进行 DNA 存储还为时过早,但干燥存储为冷冻样本提供了额外的保护,以防冷冻机故障导致样本降解。这三种 DNA 保存形式都可以在条形码设施之间运输干燥的 DNA 和 PCR 产物。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/26a5/3910151/9e5052325207/men0013-0890-f1.jpg

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