DNA extraction from honey bee intestinal bacteria and pollen: The mite Varroa destructor is largely responsible for the catastrophic reduction inviable colonies of the Dark Northern European honey bee Apis mellifera mellifera. Furthermore, introgression from imported southern sub-species is thought to havealtered the genetic integrity of this bee to the point where it is now consideredextinct across most of Europe, but Ireland remains a stronghold (Hassett et al. ,2018,Henriques et al.)
Extraction of Nucleic Acids from Sugar Beet Leaves: Within the framework of a TILLING project (Targeting Induced Local Lesions In Genomes) nucleic acids from leaf material are to be isolated from potential mutants. First a representative range of EMS (Ethyl methane sulfonate)-mutants is produced from 2000-5000 M2 plants and identified through TILLING. TILLING technology is a new and very effective method of reverse genetics for the production and identification of loss/gain of function-mutations of commercially valuable genes without genetic modification.
Perchloric Acid Extraction of Leaves for Measurement of Starch and Soluble Metabolites: The harvested plant material (up to about 300 mg) is pre-weighed into 5ml polycarbonate vials (SPEX SamplePrep cat no. 2240- PC) suitable for use in Geno/Grinder® instrument. The tubes are then capped and placed immediately into liquid nitrogen (the time from harvest to freezing should be as short as possible) NB: Plant material can be harvested and frozen in any cryo-vial, then transferred to the Geno/Grinder tubes just before grinding if necessary.
RNA Extraction from Aspergillus parasiticus mycelium: Introduction Recent genomic efforts on toxigenic and nontoxigenic Aspergillus species have advanced our understanding of the biology and genetics of these filamentous fungi. However, it is clear that these complex experiments suffer greatly from the variability in the quality of RNA between each replicate and it is critical to establish a platform to isolate high quality RNA for use in both microarray and qRT-PCR.
High Throughput Disruption of Yeast in a 96-Well Format: The wealth of information generated from years of biochemical, genetic, and molecular analyses has made yeasts both model biological systems and tools for biopharmaceutical scientists. Consequently, yeast are a popular host for gene expression studies and for the production of recombinant proteins. Though many yeast species are in use, including Pichia, Hansenula, and Debaryomyces, the most popular yeast continues to be Saccharomyces.
Extraction of RNA/cDNA and Genomic DNA from Tissue with Real-Time PCR: Fresh samples of animal tissue were collected, trimmed to approximately 50 – 100 mg of wet weight, snap-frozen in liquid nitrogen, and stored at –80˚C. The animals were human, dog, cat, mouse, cow, and horse, as well as fish and clams; see Table 1. Before DNA and/or RNA extraction, the tissues were transferred frozen to a deep-well titre plate standing on a block of dry ice.
Comparison of Pesticide Extraction in Agricultural Products Using a Manual Shaking Method and Mechanical Mixing with the Geno/Grinder®: Since its introduction in 2003 by Anastassiades and Lehotay et al, the QuEChERS method1 (quick, easy, cheap, effective, rugged, safe) has proven to be effective and convenient for analysis of multiple pesticides in agricultural products. Increasing concern over the health effects of residual pesticides on fruits and vegetables has led to increased testing of these products to determine the levels of pesticides on produce when it goes to market. The QuEChERS method has allowed analysts to process a greater number of samples in a shorter period of time than with previous methods.
Quick DNA Extraction from Rice Seed (Wet): DNA extractions can be a very time consuming and tedious process. Finding a quick method in which DNA could be extracted and used for PCR is essential. Described below is a quick “dirty” method that produces a high enough concentration of DNA that can be used for PCR.
Analysis of Neonicotinoids in Honey by QuEChERS and UHPLC-MS/MS: Neonicotinoids are a relatively new class of insecticide that were introduced as an alternative to organophosphate, carbamate and pyrethroid insecticides. Their novel mode of action works by irreversibly binding to nicotinic acetylcholine receptors, resulting in paralysis and death of insects. Since their introduction in the 1990s the neonicotinoids have been used extensively in crop protection.
Using a QuEChERS Approach for the Determination of Pesticide Residues in Soil: The use of pesticides in agriculture and households is widespread. To ensure food safety and prevent the unnecessary exposure of consumers to pesticides it is important to test for these residues in surveillance plans. While the greatest source of pesticide exposure comes from residues that remain in final food products, they can also be found in environmental samples such as water and soil.
Bead Beating – Introduction & Guide: Bead beating is an effective process used to disrupt a wide range of biological samples. It is achieved by rapidly agitating samples with grinding media (balls or beads) in a bead beater. Samples can be processed with or without buffer or solvent at room temperature or cryogenically.
The Application of QuEChERS in the Extraction of Anabolic Steroids in Whole Blood: Anabolic steroids are drugs structurally related to the cyclic steroid ring system and behave similarly to testosterone in the body. Anabolic steroids are used therapeutically to stimulate muscle growth and appetite, induce male puberty and treat chronic wasting conditions, such as cancer and AIDS. Ergogenic uses of anabolic steroids include bodybuilding, sport doping, and animal fattening.
Determination of 11-nor-9-Carboxy-THS in Human Urine by QuEChERS and LC-MS/MS: 11-nor-9-Carboxy-THC, also known as THCA or carboxy-THC, is the main secondary metabolite of THC (the active component of marijuana) formed in the human body. THCA is excreted in urine in the form of glucuronide conjugates. THCAis not psychoactive but has a long half-life up to several days or even weeks in very heavy users, thus determination of THCA in urine plays an important role in confirmation of marijuana consumption.
Determination of Pesticides in Bananas by QuEChERS and LC-MS/MS: This application describes a simple, fast, and cost-effective method for the determination of multi-class pesticides in bananas including one of the most difficult compounds, pymetrozine. The method employs the AOAC version of the QuEChERS procedure, in which 15 g of the homogenized banana sample is hydrated with 5 mL of reagent water to give a sample with > 80% water content. The hydrated sample is extracted by 15 mL of acetonitrile (MeCN) with 1% (v/v) acetic acid (HAc), followed by the addition of 6 g anhydrous magnesium sulfate (MgSO4) and 1.
Removal of Purple Pigmentation from Cannabis using QuEChERS Extraction, ChloroFiltr® dSPE Clean-up and LC-MS/MS: Cannabis testing laboratories have the challenge of removing a variety of unwanted matrix components from plant material prior to running extracts on their LC-MS/MS or GC-MS. The complexity of the cannabis plant presents additional analytical challenges that do not need to be accounted for in other agricultural products. Up to a third of the overall mass of cannabis seed, half of usable flower, and nearly all of extracts can be contributed to essential oils such as terpenes, flavonoids and actual cannabinoid content.
Comparison of Methods for the Isolation of DNA from Soybeans: The isolation of nucleic acids from intact seeds requires mechanically disrupting the seed followed by the extraction and subsequent purification of the nucleic acid. The mechanical disruption is often performed manually with a mortar and pestle, an approach that is not practical for high throughput screening of seeds as manual grinding is slow and reuse of mortar and pestles may lead to cross-contamination. Alternatively, nucleic acids can be isolated from seeds in a microwell plate format using a ball mill that mechanically disrupts the seeds.
Extraction of Cannabinoids in Marijuana and Edibles by QuEChERS: Medical marijuana has been legalized in multiple states across the USA. As a result, many testing labs are seeking fast and reliable analytical methods to determine the cannabinoid potency in marijuana and cannabis infused foods (more informally known as edibles). This application utilizes the advantages of the QuEChERS technique to extract cannabinoids in marijuana and cannabis containing foods.
Determination of Mycotoxin Residues by LC-MS/MS Featuring Two Alternate Sample Extraction Procedures: Mycotoxins are toxic natural metabolites produced by several species of fungi on agricultural commodities in the field or during storage. To date more than 300 mycotoxins, possessing varying degrees of toxicity, have been identified, although only a relatively few of these are widely accepted as presenting a significant food or animal feed safety risk . Mycotoxins are chemically stable and cannot be destroyed during food processing and heat treatment, thus, monitoring these compounds in food is an important health, agricultural production, food processing and trade concern.
Pesticide Residue Analysis in Whole milk by QuEChERS and LC-MS/MS: This application describes a cost-effective and easy to use method of the determination of pesticide residues in whole milk. This method employs the AOAC version of QuEChERS. This procedure provides better analytical results than either the original or EN versions of the QuEChERS procedure in extracting a few sensitive pesticides, such as pymetrozine and hexazinone (Velpar).
Multiresidue Analysis of Veterinary Drugs in Bovine Liver by LC-MS-MS: Agilent Bond Elut Enhanced Matrix Removal—Lipid (EMR—Lipid) is the next generation of sample preparation product and is available for convenient dispersive solid phase extraction (dSPE). The material is highly selective towards coextracted matrix, especially from fatty samples (fat content > 5%) without negatively impacting analyte recovery. This study demonstrates the application of this novel product for the analysis of 30 representative veterinary drugs in bovine liver.