Ethanol preservation and pretreatments facilitate quality DNA extractions in recalcitrant plant species

• Published in: Applications in plant sciences Vol. 11; no. 3; pp. e11519 - n/a
• Main Authors: Johnson, Gabriel, Canty, Steven W. J., Lichter‐Marck, Isaac H., Wagner, Warren, Wen, Jun
• Format: Journal Article
• Language: English
• Published: United States John Wiley & Sons, Inc 01.05.2023; John Wiley and Sons Inc; Wiley
• Subjects: Application; cetyltrimethylammonium bromide; Deoxyribonucleic acid; digestion; DNA; Enzymes; Ethanol; Flowers & plants; Freezing; herbaria; herbarium; Leaves; liquid nitrogen; Lysis; mangrove; Plant sciences; plant tissues; Preservation; proteinases; recalcitrant species; Rhizophora mangle; Sorbitol; tissue preservation; Vitaceae; United States > US; herbarium; tissue preservation; Rhizophora mangle; mangrove; Vitaceae; ethanol
• ISSN: 2168-0450; 2168-0450
• DOI: 10.1002/aps3.11519

Premise The preservation of plant tissues in ethanol is conventionally viewed as problematic. Here, we show that leaf preservation in ethanol combined with proteinase digestion can provide high‐quality DNA extracts. Additionally, as a pretreatment, ethanol can facilitate DNA extraction for recalcitrant samples. Methods DNA was isolated from leaves preserved with 96% ethanol or from silica‐desiccated leaf samples and herbarium fragments that were pretreated with ethanol. DNA was extracted from herbarium tissues using a special ethanol pretreatment protocol, and these extracts were compared with those obtained using the standard cetyltrimethylammonium bromide (CTAB) method. Results DNA extracted from tissue preserved in, or pretreated with, ethanol was less fragmented than DNA from tissues without pretreatment. Adding proteinase digestion to the lysis step increased the amount of DNA obtained from the ethanol‐pretreated tissues. The combination of the ethanol pretreatment with liquid nitrogen freezing and a sorbitol wash prior to cell lysis greatly improved the quality and yield of DNA from the herbarium tissue samples. Discussion This study critically reevaluates the consequences of ethanol for plant tissue preservation and expands the utility of pretreatment methods for molecular and phylogenomic studies.