The Development of a Rapid and Sensitive, High-Through-Put Protocol for RNA:DNA Ratio Analysis

Kaplan, Lisa A. E.; Leamon, John H.; Crivello, Joseph F.

doi:10.1577/1548-8667(2001)013<0276:tdoara>2.0.co;2

Cited by 11 publications

(10 citation statements)

References 8 publications

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“…Nucleic acid quantification procedure used was based on the fluorometric method described by Kaplan, Leamon and Crivello (). Lyophilized caudal peduncles from 5 larvae were pooled, weighed, homogenized and digested for 2 h at 55°C in a solution containing 0.1 M NaCl, 0.2% sodium dodecylsulfate and 10 mg mL −1 proteinase K. Following digestion, samples were centrifuged (13 000 g , 10 min).…”

Section: Methodsmentioning

confidence: 99%

Stimulatory effect of thyroid hormones improves larval development and reproductive performance in alligator gar (Atractosteus spatula) and spotted gar (Lepisosteus oculatus)

et al. 2013

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Development of alligator gar (Atractosteus spatula) and spotted gar (Lepisosteus oculatus) larvae was evaluated after exposure of embryos to thyroid hormones (TH) by two different methods of administration. For the first method, alligator gar embryos were placed in a water bath with triiodothyronine (T3; 1 ppm for 2 h), and control treatment embryos were placed in a water bath without T3. For the second method, spotted gar adult males and females were injected with different TH during induced spawning: a group was injected with T3 [20 mg kg−1 body weight (BW)], a second group with thyroxine (T4; 20 mg kg−1 BW), a third group with thyroid‐stimulating hormone (TSH; 4 IU kg−1BW) and a control group with dimethyl sulfoxide. Both methods resulted in increases in the concentrations of T3 and T4 in embryos of both species. An increase in the alkaline enzymatic activity of TH‐treated larvae was observed as well as an increase in the RNA:DNA, RNA:Dry weight and DNA:Dry weight ratios at hatching. Some positive effects of TH application were: a significant increase in the hatching rate of spotted gar larvae from TH‐injected parents and a higher percentage of successful spawns. Evenness of larval sizes and a higher survival rate were observed in alligator gar larvae. Snout development was accelerated by TH treatment in both species.

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Section: Methodsmentioning

confidence: 99%

Stimulatory effect of thyroid hormones improves larval development and reproductive performance in alligator gar (Atractosteus spatula) and spotted gar (Lepisosteus oculatus)

et al. 2013

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“…Frozen or living individual adult female Epischura were homogenized in 25 µL of 50 mM TE buffer, pH 8.0 containing 60 mAU/ml proteinase K (Qiagen). Proteinase K, rather than N-laurylsarcosine extraction that follows Kaplan et al (2001) protocol was chosen due to greater RNA stability during the 20-40 min window following extraction (ESM Figure S2), in contrast to McGinty et al, 2008. Homogenates were vortexed briefly and incubated for 20 min in a 54°C water bath, with periodic vortexing.…”

Section: Rna and Dna Extraction And Determinationmentioning

confidence: 99%

A capital breeder in a heterogeneous environment: Lipid reserves and RNA:DNA ratio in Lake Baikal's endemic Epischura

Bowman

Kondrateva

Silow

et al. 2017

Journal of Great Lakes Research

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Lake Baikal (Siberia, Russia) is the oldest, the deepest and the most voluminous lake on Earth. During the last century, Baikal has experienced gradual increases in temperature but remains highly oligotrophic with increases in primary productivity limited to the warmest parts of the lake. Using whole body RNA:DNA ratio as an indicator of metabolic rate, we demonstrate that the key primary consumer in Baikal's plankton, Epischura baikalensis (Copepoda: Calanoida), is a capital breeder that relies on lipid storage to maintain productivity. In individuals from nature, the RNA:DNA ratio correlated with lipid content in samples from cold, low phytoplankton density locations from Baikal's North and Central basins, but not in samples from warmer South basin and Maloe More strait. Lipid reserves, both visually assessed and measured by Nile Red fluorescence, correlated positively with phytoplankton density. In laboratory experiments the RNA:DNA ratio responded to starvation and temperature in non-reproducing, low lipid storage females, but not in individuals with developed ovaries or high lipid storage. This indicates that, unlike many other zooplankton crustaceans, E. baikalensis uses resource storage to support current reproduction, which buffers the dependency of metabolic rate on current feeding conditions. We discuss possible effects of such buffering on E. baikalensis competition with non-endemic, largely income-feeding zooplankton species whose frequency is currently increasing in Baikal pelagia.

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“…Fourth, for RNA/DNA determinations, results often depend on the technique used to determine nucleic acid content, in part because RNA is much more sensitive to enzymatic degradation than DNA. Recent advances in nucleic acid quantification techniques have minimized this final problem (84,85), but the other points remain considerations for applying these techniques to ecological studies.…”

Section: Rna/dna As Indicator Of Conditionmentioning

confidence: 99%

Biochemical Indicators of Stress and Metabolism: Applications for Marine Ecological Studies

Dahlhoff

2004

Annu. Rev. Physiol.

259

175

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Studies investigating the effects of temperature, food availability, or other physical factors on the physiology of marine animals have led to the development of biochemical indicators of growth rate, metabolic condition, and physiological stress. Measurements of metabolic enzyme activity and RNA/DNA have been especially valuable as indicators of condition in studies of marine invertebrates and fishes, groups for which accurate determination of field metabolic rates is difficult. Properly calibrated and applied, biochemical indicators have been successfully used in studies of rocky intertidal ecology, where two decades of experimentation have generated rigorous, testable models for determining the relative influences of biotic and abiotic factors on species distribution, abundance, and interaction. Biochemical indicators of condition and metabolic activity (metabolic enzymes, RNA/DNA) have been used to test nutrient-productivity models by demonstrating tight linkages between nearshore oceanographic processes (such as upwelling) and benthic rocky intertidal ecosystems. Indices of condition and heat stress (heat shock proteins, or Hsps) have begun to be used to test environmental stress models by comparing condition, activity, and Hsp expression of key rocky intertidal predator and prey species. Using biochemical indicators of condition and stress in natural systems holds great promise for understanding mechanisms by which organisms respond to rapid environmental change.

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The Development of a Rapid and Sensitive, High-Through-Put Protocol for RNA:DNA Ratio Analysis

Cited by 11 publications

References 8 publications

Stimulatory effect of thyroid hormones improves larval development and reproductive performance in alligator gar (Atractosteus spatula) and spotted gar (Lepisosteus oculatus)

Stimulatory effect of thyroid hormones improves larval development and reproductive performance in alligator gar (Atractosteus spatula) and spotted gar (Lepisosteus oculatus)

A capital breeder in a heterogeneous environment: Lipid reserves and RNA:DNA ratio in Lake Baikal's endemic Epischura

Biochemical Indicators of Stress and Metabolism: Applications for Marine Ecological Studies

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