Storage effects on genomic DNA in rolled and mature coca leaves

Abstract

Rolled and mature leaf tissue was harvested from Erythroxylum coca var. coca Lam. (coca) to determine a method for storage that would maintain DNA with high quality and content up to 50 days. Harvesting coca leaf tissue under Andean field conditions often requires storage from 3 to 10 days before extraction where tissue integrity is lost. All samples of rolled and mature coca leaf tissue were harvested and separately stored fresh in RNAlaterTM for 50 days at 4, 20, and 23C, while similar samples were air-dried for 72 h at 23C or ovendried for 72 h at 40C after storage, before extraction. Triplicate samples of each tissue type were extracted for DNA at 10-day intervals and showed that DNA integrity and content were preserved in leaf tissue stored at 4 and 20C for 50 days. Rolled and mature leaf tissue stored at 4, 20, and 23C showed insignificant degradation of DNA after 10 days, and by day 50, only leaf tissue stored at 4 and 20C had not significantly degraded. All air- and oven-dried leaf tissue extracts showed degradation upon drying (day 0) and continuous degradation up to day 50, despite storage conditions. Amplified fragment length polymorphism analysis of DNA from rolled and mature leaf tissue of coca stored at 4 and 20C for 0, 10, and 50 days showed that DNA integrity and content were preserved. We recommend that freshly harvested rolled or mature coca leaf tissue be stored at 4, 20, and 23C for 10 days after harvest, and if a longer storage is required, then store at 4 or 20C.

Effects of preservation and storage of microcrustaceans in RNAlateron RNA and DNA degradation

“…The degradation of RNA as a result of freezing has been indicated previously by the formation of breakdown products of 28S rRNA extracted from snapfrozen autopsy tissues (Ross et al 1992). Similar effects have also been reported for DNA with both statistically significant (Pesaro et al 2003) and not significant (Johnson et al 2003) degradation attributed to freeze-thaw stress.…”

“…RNAlater ® (Ambion) is a storage reagent that stabilizes and protects cellular RNA in intact unfrozen samples by rapidly penetrating fresh tissue and deactivating nucleases. The reagent is also applicable for DNA preservation (Gorokhova and Kyle 2002;Johnson et al 2003). According to the general directions of the manufacturer, samples can be stored in RNAlater for up to 1 day at 37°C, for up to a week at room temperature, for a month or more at 4°C, or stored indefinitely at -20°C without nucleic acid degradation (Ambion 2001).…”

Spatial variation in RNA:DNA ratios of Diporeia spp. in the Great Lakes region

“…Upon collection, live animals were gently screened from the sediments and individuals immediately placed in vials with 1.0 mL RNAlater®, a supersaturated salt solution that preserves nucleic acids. Several past studies have successfully relied on RNAlater® to preserve nucleic acids of various experimental and in situ collected animals (Gorokhova and Kyle, 2002;Gorokhova, 2003;Johnson et al, 2003;Höök et al, 2008).…”

OPTIMIZATION OF DNA EXTRACTION FROM BROWN ALGAE (PHAEOPHYCEAE) BASED ON A COMMERCIAL KIT1

“…It appears that both methods of preservation might not be ideal procedures, because in either case, drying was rather slow and could have led to stresses triggering DNA degradation. Generally, for animal samples, once the tissues are dried, DNA is fairly stable; however, in plants and algae, DNA degradation frequently occurs, even in dried tissues (Johnson et al. 2003, Ribeiro and Lovato 2007).…”