Reliable detection and identification of genetically modified maize, soybean, and canola by multiplex PCR analysis

Multiplex PCR procedures were developed for simultaneously detecting multiple target sequences in genetically modified (GM) soybean (Roundup Ready), maize (event 176, Bt11, Mon810, T14/25), and canola (GT73, HCN92/28, MS8/RF3, Oxy 235). Internal control targets (invertase gene in corn, lectin and beta-actin genes in soybean, and cruciferin gene in canola) were included as appropriate to assess the efficiency of all reactions, thereby eliminating any false negatives. Primer combinations that allowed the identification of specific lines were used. In one system of identification, simultaneous amplification profiling (SAP), rather than target specific detection, was used for the identification of four GM maize lines. SAP is simple and has the potential to identify both approved and nonapproved GM lines. The template concentration was identified as a critical factor affecting efficient multiplex PCRs. In canola, 75 ng of DNA template was more effective than 50 ng of DNA for the simultaneous amplification of all targets in a reaction volume of 25 microL. Reliable identification of GM canola was achieved at a DNA concentration of 3 ng/microL, and at 0.1% for GM soybean, indicating high levels of sensitivity. Nonspecific amplification was utilized in this study as a tool for specific and reliable identification of one line of GM maize. The primer cry1A 4-3' (antisense primer) recognizes two sites on the DNA template extracted from GM transgenic maize containing event 176 (European corn borer resistant), resulting in the amplification of products of 152 bp (expected) and 485 bp (unexpected). The latter fragment was sequenced and confirmed to be Cry1A specific. The systems described herein represent simple, accurate, and sensitive GMO detection methods in which only one reaction is necessary to detect multiple GM target sequences that can be reliably used for the identification of specific lines of GMOs.     

Diversified cropping systems in semiarid Montana: Nitrogen use during drought

Improved nitrogen use efficiency would be beneficial to agroecosystem sustainability in the northern Great Plains of the USA. The most common rotation in the northern Great Plains is fallow–spring wheat. Tillage during fallow periods controls weeds, which otherwise would use substantial amounts of water and available nitrogen, decreasing the efficiency of fallow. Chemical fallow and zero tillage systems improve soil water conservation, and may improve nitrogen availability to subsequent crops. We conducted a field trial from 1998 through 2003 comparing nitrogen uptake and nitrogen use efficiency of crops in nine rotations under two tillage systems, conventional and no-till. All rotations included spring wheat, two rotations included field pea, while lentil, chickpea, yellow mustard, sunflower, and safflower were present in single rotations with wheat. Growing season precipitation was below average in 3 of 4 years, resulting in substantial drought stress to crops not following fallow. In general, rotation had a greater influence on spring wheat nitrogen accumulation and use efficiency than did tillage system. Spring wheat following fallow had substantially higher N accumulation in seed and biomass, N harvest index, and superior nitrogen use efficiency than wheat following pea, lentil, chickpea, yellow mustard, or wheat. Preplant nitrate-N varied widely among years and rotations, but overall, conventional tillage resulted in 9 kg ha⁻¹ more nitrate-N (0–60 cm) for spring wheat than did zero tillage. However, zero tillage spring wheat averaged 11 kg ha⁻¹ more N in biomass than wheat in conventional tillage. Nitrogen accumulation in pea seed, 45 kg ha⁻¹, was superior to that of all alternate crops and spring wheat, 17 and 23 kg ha⁻¹, respectively. Chickpea, lentil, yellow mustard, safflower, and sunflower did not perform well and were not adapted to this region during periods of below average precipitation. During periods of drought, field pea and wheat following fallow had greater nitrogen use efficiency than recropped wheat or other pulse and oilseed crops.     

Wireless lysimeters for real-time online soil water monitoring

Identification of drainage water allows assessing the effectiveness of water management. Passive capillary wick-type lysimeters (PCAPs) were used to monitor water flux leached below the root zone under an irrigated cropping system. Wireless lysimeters were developed for web-based real-time online monitoring of drainage water using a distributed wireless sensor network (WSN). Twelve PCAP sensing stations were installed across the field at 90 cm below the soil surface, and each station measured the amount of drainage water using two tipping buckets mounted in the lysimeter and continually monitored soil water contents using two soil moisture sensors installed above the lysimeter. A weather station was included in the WSN to measure micrometeorological field conditions. All in-field sensory data were periodically sampled and wirelessly transmitted to a base station that was bridged to a web server for broadcasting the data on the internet. Communication signals from the in-field sensing stations to the base station were successfully interfaced using low-cost Bluetooth wireless radio communication. Field experiments resulted in high correlation between estimated and actual drainage with r ² = 0.95 and confirmed a reliable wireless communication throughout the growing season. A web-linked WSN system provided convenient remote online access to monitor drainage water flux and field conditions without the need for costly time-consuming supportive operations.     

An evaluation of a steerable sidescan sonar for surveys of near-surface fish

A towed 330 kHz sector scanning sonar was evaluated for surveys of near-surface migratory salmonids (Oncorhynchus sp.) and other fish in the Strait of Georgia near the mouth of the Fraser River near Vancouver, BC (Canada). Detection of large near-surface fish presents a serious challenge to conventional net trawls or echo-sounder surveys due to vessel avoidance behaviour by the fish. The sonar was towed beside a research vessel on a custom-built towfish at a nominal depth of 18 m. The optimum configuration was determined to be with the sonar scanning a 30° vertical sector oriented perpendicular to the tow direction, at a ping rate of 5 Hz and a sector scan period of 5 s. Fish were detected up to a range of 100 m, limited by systemic noise and backscattered reverberation from the sea surface. Some evidence of vessel-avoidance behaviour by the fish was observed. Near surface temperature and salinity gradients are shown to induce downwards refraction of the sonar beam, resulting in biases in apparent depth and target strength of the fish. A simple model of the sonar performance in this scanning mode, including beam-pattern effects, is used to generate curves of fish target detectability and beam-pattern induced bias as a function of range and fish target strength. After correction for refraction effects, beam-pattern bias, and detectability, the survey results show reasonable agreement with net trawl and riverine escapement data.     

Efficacy of starter N fertilizer and rhizobia inoculant in dry pea (Pisum sativum Linn.) production in a semi-arid temperate environment

ABSTRACT

Uncertainties exist about the importance of rhizobia inoculant and starter nitrogen (N) application in dry pea (Pisum sativum L.) production. Three field experiments were conducted to evaluate how rhizobia inoculant and starter N fertilizer affect pea seed yield and protein concentration in a semi-arid environment in central Montana. Commercial rhizobia inoculant was mixed with seed prior to planting at the manufacturer’s recommended rate. Starter N fertilizers were applied into the same furrow as seed at 0, 22, 44 and 88 kg ha^?1 as urea, slow-release polymer-coated N fertilizer (ESN), and a combination of both. The application of rhizobia inoculant had no or a very small beneficial effect on pea yield in lands with a previous history of peas. In a land without pea history, application of rhizobia increased pea seed yield by 16%. The positive effect of starter N was only pronounced when initial soil N was low (≤ 10 kg ha^?1 nitrate-nitrogen), which increased net return by up to US$ 42 ha^?1. In this condition, application of slow-release N outperformed urea. However, application of starter N (especially with urea) had a negative effect on pea establishment, vigor and seed yield when soil initial N was high (≥ 44 kg ha^?1 NO₃-N). The results indicate that the rate, placement and form of the starter N must be optimized to benefit pea yield and protein without detrimental effects on germination and nodulation. Moreover, application of starter N must be guided by the soil nitrate content.     

Aboriginal Precedent for Active Management of Sagebrush-Perennial Grass Communities in the Great Basin

Until recently, most contemporary ecologists have ignored or diminished anecdotal historical accounts and anthropologists' reports about aboriginal fire in the Great Basin. Literature review shows that Indians practiced regular use of fire for many purposes, including the obvious reasons of increasing the availability of desired plants, maintaining habitats for animals used as food, and driving game during hunts. Historical accounts of prehistoric anthropogenic firing, inferences from fire-scar data, and data regarding annual production capability of representative sagebrush (Artemisia spp.)-perennial grass ecological sites indicate that prehistoric conditions were neither fuel- nor ignition-limited. According to many sources, this “active management” by Indians was widespread, significant, and more common than lightning-caused fires, resulting in mosaic vegetation patterns that subsequently moderated the behavior of “natural fires.” This interaction between Indian-burning and lightning fires may have strongly influenced the pre-Euro-American settlement vegetation of the Great Basin. At the very least, the landscape was a patchwork of areas altered by aboriginal people and areas shaped primarily by bio-physical processes. Based on this prehistoric precedent, current historically unprecedented conditions (fuel load and exotic weed invasion threats), and predicted climate change, contemporary active management of sagebrush-perennial grass communities is paramount. Restoration measures should be scientifically based and tailored to achieve ecological resilience and functionality in specific sites. Prescribed fire is not always ecologically appropriate or judicious, especially in Wyoming big sagebrush (A. tridentata spp. wyomingensis) communities, so managers should consider using other alternatives where an intentional low severity distubance is deemed necessary. Properly planned active management would disrupt fuel continuity for lighthning fires, ensure ecological process and successional integrity, and benefit multiple uses on a landscape scale.     

Challenges for the Kasatoshi volcano hypothesis as the cause of a large return of sockeye salmon (Oncorhynchus nerka) to the Fraser River in 2010

In August 2008 the Kasatoshi volcano in the Aleutian archipelago erupted. Prevailing winds carried volcanic ash to the Gulf of Alaska, where its soluble iron dissolved and initiated a widespread phytoplankton bloom. Two years later, the abundance of sockeye salmon (Oncorhynchus nerka) returning to spawn in the Fraser River was larger than any observed since 1913. Kasatoshi's influence on growth and survival has been proposed as the ultimate cause of the abundant return. However, when relevant data are brought to bear on the hypothesis, it shows that: (1) survival of the abundant sockeye salmon cohort was unremarkable when compared with the historical record; (2) without an accompanying volcano, survival of the returns in 2011 was about the same as in 2010; (3) parental abundance that spawned the abundant return in 2010 was the sixth largest since 1948; (4) during their first summer at sea, sockeye salmon are not known to migrate in the offshore region where the anomalous chlorophyll bloom occurred; (5) an older cohort that was likely exposed to the chlorophyll bloom had the second lowest productivity on record when it returned in 2009; (6) immature sockeye salmon do not feed on diatoms, so any trophodynamic benefit derived from excess chlorophyll in mid‐to late August 2008 would have to have found a rapid trophic pathway to sockeye salmon on the continental shelf; and (7) no other populations of sockeye salmon or other species of salmon that are known to rear in the same region had unexpectedly high returns.     

Evaluation of poultry fat and blends of poultry fat with cold‐pressed flaxseed oil as supplemental dietary lipid sources for juvenile sablefish (Anoplopoma fimbria)

The objective of this study was to access the suitability of using poultry fat (PF) or blends of PF with flaxseed oil (FO) to replace 75% of the supplemental anchovy oil (AO) in the diet of juvenile sablefish (Anoplopoma fimbria), a relatively new marine species to aquaculture. Sablefish were fed one of four diets twice daily to satiation for 15 weeks. The test diets were identical in composition except for the source of supplemental lipid which was either 100% AO (100AO), or had 75% of the supplemental AO replaced with 50% FO:25% PF, 25% FO:50% PF or 75% PF. Sablefish growth parameters, whole body and fillet proximate constituent concentrations, and apparent digestibility coefficients were uninfluenced by diet treatment. There were also no adverse effects of the diet treatments on fish health, as determined from analysis of various haematological and innate immunological parameters. Terminal fillet fatty acid compositions generally reflected the dietary fatty acid compositions. Results indicated that PF or blends of PF and FO may comprise 75% of the supplemental lipid in a grower diet for sablefish and are an economic alternative to AO while still providing humans with a rich dietary source of highly unsaturated fatty acids.     

Oceanic and atmospheric extremes motivate a new hypothesis for variable marine survival of Fraser River sockeye salmon

In spite of a relatively optimistic pre‐season forecast, the total return of adult sockeye salmon (Oncorhynchus nerka) to the Fraser River (British Columbia, Canada) in 2009 was the lowest recorded since quantitative records began in the late 1940s. A plausible mechanism is proposed that links a sequence of extreme oceanic and climatic events to poor marine survival. It began with record‐setting snow packs in the coastal mountain range during the winter of 2007 that led to the development of unprecedented oceanographic conditions in the spring of 2007 from Queen Charlotte Strait in central British Columbia to Southeast Alaska. When combined with equally extreme atmospheric anomalies in the region in the spring of 2007, with a winter wind regime persisting through July, a coastal surface ocean with characteristics that are known to be associated with lower marine survival was established. Most of the sockeye salmon that were expected to return to the Fraser River as adults in 2009 passed through this atypical ocean as juveniles on their migration to the open ocean in 2007. A trophic gauntlet hypothesis is proposed as a new paradigm to describe the oceanic environment faced by sockeye salmon after they emigrate northward from the Strait of Georgia. The hypothesis identifies a new type of high nutrient low chlorophyll region that can explain how oceanographic extremes at critical locations along the migration route beyond the Strait of Georgia can reduce marine survival in some years.     

Real-time polymerase chain reaction (PCR) quantitative detection of Brassica napus using a locked nucleic acid TaqMan probe

Several countries have introduced mandatory labeling requirements on foods derived from genetically modified organisms. Real-time quantitative Polymerase Chain Reaction (PCR) has quickly become the method of choice in support of these regulations and requires the development of separate PCR assays targeting the transgenic sequence as well as a specific endogenous gene sequence. To develop a Brassica napus-specific PCR assay, partial sequences of the acetyl-CoA carboxylase BnACCg8 gene from B. napus and the closely related Brassica rapa were determined and compared, and a region of unique nucleotide sequence was identified. Universal amplification primers were designed to either side of this region, and a locked nucleic acid TaqMan probe was designed to the B. napus-specific sequence. Evaluation of this primer/probe combination indicated a high level of specificity to B. napus: no amplification signal was observed with any other species tested, including five closely related Brassica species. The method was assayed with 14 different B. napus cultivars, and comparable amplification curves were consistently obtained for all. The assay was highly sensitive, with a limit of detection between 1 and 10 haploid copies. Practically, the method was demonstrated to be effective for the detection of processed food samples and for the quantification of Roundup Ready canola content in mixed samples.