一种适合野外使用的被子植物分子标本干燥方式

doi:10.17521/cjpe.2016.0322

摘要/Abstract

摘要：

该研究通过比较不同干燥方式对被子植物分子标本的影响, 试图得到一种野外采集过程中可以替代硅胶干燥法的更为便捷的植物分子标本的干燥方法。选取40、80和150 ℃烘干和吸水纸压制干燥、硅胶干燥法对日本晚樱(Prunus serrulata var. lannesiana)和山麦冬(Liriope spicata)两种植物的新鲜叶片进行干燥处理, 提取各种处理样品的DNA, 并将DNA进行电泳检测、分光光度计检测及PCR扩增, 以此评价不同的干燥方式对植物基因组DNA的影响。分光光度计检测及总DNA电泳结果显示, 经40 ℃烘干或硅胶干燥处理的样品总DNA浓度及长片段DNA浓度较其他干燥方式高; PCR产物浓度统计学分析显示, 40 ℃烘干处理的样品PCR产物浓度高于其他干燥方式。基于以上结果, 建议在野外采集被子植物分子标本时, 使用40 ℃烘干干燥法对分子标本进行干燥处理, 避免分子标本快速降解以及硅胶干燥法的污染问题, 同时可节省携带、更换大量硅胶材料所耗费的人力。

关键词: 被子植物, 野外, 分子标本, DNA, 烘干, 硅胶

Abstract:

Aims This study aims to identify a more convenient drying method for obtaining molecular specimen of angiosperms in the field than the conventional silica gel drying method. Methods The leaves of Prunus serrulata var. lannesiana and Liriope spicata were dried under temperatures of 150 °C, 80 °C, 40 °C as well as under natural conditions, and by the silica gel drying method, respectively. The DNA extracts of various specimens were then analyzed using techniques of spectrophotometer detection, electrophoresis and PCR to evaluate the impacts of different drying treatments to the genomic DNA of testing plants. Important findings The concentrations of total DNA were higher for the specimens dried at 40 °C treatment and by the silica gel drying method than other treatments when assessed by the techniques of spectrophotometer detection and electrophoresis. The concentration of PCR products was highest in the specimens dried at 40 °C. Based on the results, the 40 °C drying can be recommended for obtaining molecular specimens of angiosperms because of its minimum degree of degradation, for convenience of operation and avoiding carrying large amounts of silica gel in field investigations.

Key words: angiosperms, field, molecular specimen, DNA, drying, silica gel

沈风娇, 任倩倩, 董琦, 朱丽, 张建芳, 杨婧, 张冉, 梁红柱, 赵建成, 石硕. 一种适合野外使用的被子植物分子标本干燥方式. 植物生态学报, 2017, 41(7): 787-794. DOI: 10.17521/cjpe.2016.0322

Feng-Jiao SHEN, Qian-Qian REN, Qi DONG, Li ZHU, Jian-Fang ZHANG, Jing YANG, Ran ZHANG, Hong-Zhu LIANG, Jian-Cheng ZHAO, Shuo SHI. A new angiosperms molecular specimen treatment method for field use. Chinese Journal of Plant Ecology, 2017, 41(7): 787-794. DOI: 10.17521/cjpe.2016.0322

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链接本文: https://www.plant-ecology.com/CN/10.17521/cjpe.2016.0322

https://www.plant-ecology.com/CN/Y2017/V41/I7/787

图/表 5

表1 日本晚樱和山麦冬分子标本经不同干燥处理后DNA纯度、浓度及PCR产物浓度的比较(平均值±标准偏差)

Table 1 Comparisons of DNA purity, concentration and the concentration of PCR products in the specimens of Prunus serrulata var. lannesiana and Liriope spicata obtained with different drying methods (mean ± SD)

干燥方式 Drying method	日本晚樱 Prunus serrulata var. lannesiana			山麦冬 Liriope spicata
干燥方式 Drying method	OD260/280	DNA (ng·μL^-1)	PCR (ng·μL^-1)	OD260/280	DNA (ng·μL^-1)	PCR (ng·μL^-1)
150	1.83 ± 0.17^a	451.24 ± 150.91^a	13.94 ± 3.97^c	2.03 ± 0.04^a	211.43 ± 64.31^a	0.74 ± 2.30^d
80	1.80 ± 0.13^a	376.13 ± 121.04^a	34.48 ± 9.42^b	1.83 ± 0.16^b	80.64 ± 48.16^c	16.51 ± 17.49^c
40	1.74 ± 0.16^ab	470.44 ± 228.50^a	49.07 ± 4.83^a	1.90 ± 0.08^ab	241.41 ± 88.07^a	57.64 ± 6.51^a
Y	1.73 ± 0.09^ab	291.64 ± 90.77^b	38.38 ± 4.75^b	1.93 ± 0.14^ab	160.01 ± 22.95^b	46.98 ± 8.22^b
S	1.64 ± 0.10^b	432.59 ± 167.67^a	39.95 ± 5.82^b	1.90 ± 0.24^ab	252.33 ± 61.74^a	43.19 ± 8.72^b

图1 5种不同干燥方式处理的日本晚樱(R)总DNA电泳图。150, 150 ℃烘干处理; 80, 80 ℃烘干处理; 40, 40 ℃烘干处理; Y, 吸水纸压制干燥; S, 硅胶干燥; 2K, DNA ladder中最长的DNA片段为2 000 bp。

Fig. 1 Genomic DNA in specimens of Prunus serrulata var. lannesiana (R) obtained with five different drying methods. 150, drying at 150 °C; 80, drying at 80 °C; 40, drying at 40 °C; Y, natural drying; S, silica gel drying; 2K, 2 kb plus DNA ladder.

图2 5种不同干燥方式处理的山麦冬(M)总DNA电泳图。150, 150 ℃烘干处理; 80, 80 ℃烘干处理; 40, 40 ℃烘干处理; Y, 吸水纸压制干燥; S, 硅胶干燥; 2K, DNA ladder中最长的DNA片段为2 000 bp。

Fig. 2 Genomic DNA in specimens of Liriope spicata (M) obtained with five different drying methods. 50, drying at 150 °C; 80, drying at 80 °C; 40, drying at 40 °C; Y, natural drying; S, silica gel drying; 2K, 2 kb plus DNA ladder.

图3 日本晚樱(R)经5种不同干燥方式处理后PCR凝胶电泳检测结果。150, 150 ℃烘干处理; 80, 80 ℃烘干处理; 40, 40 ℃烘干处理; Y, 吸水纸压制干燥; S, 硅胶干燥; 2K, DNA ladder中最长的DNA片段为2 000 bp。

Fig. 3 Agarose gel electrophoresis of PCR products in specimens of Prunus serrulata var. lannesiana (R) obtained with five different drying methods. 150, drying at 150 °C; 80, drying at 80 °C; 40, drying at 40 °C; Y, natural drying; S, silica gel drying; 2K, 2kb plus DNA ladder.

图4 山麦冬(M)经5种不同干燥方式处理后PCR凝胶电泳检测结果。150, 150 ℃烘干处理; 80, 80 ℃烘干处理; 40, 40 ℃烘干处理; Y, 吸水纸压制干燥; S, 硅胶干燥; 2K, DNA ladder中最长的DNA片段为2 000 bp。

Fig. 4 Agarose gel electrophoresis of PCR products in specimens of Liriope spicata (M) obtained with five different drying methods. 150, drying at 150 °C; 80, drying at 80 °C; 40, drying at 40 °C; Y, natural drying; S, silica gel drying; 2K, 2 kb plus DNA ladder.

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