![]() ![]() In general, the genomoviruses are classified at a species level based on their genome-wide pairwise identity with a species cutoff threshold of 78%. The family Genomoviridae is divided into nine genera ( Gemycircularvirus, Gemyduguivirus, Gemygorvirus, Gemykibivirus, Gemykolovirus, Gemykrogvirus, Gemykroznavirus, Gemytondvirus and Gemyvongvirus). Genomoviruses have an ambisense genome organization and genomes that are ~ 1.9–2.3 kb encoding a CP on the virion sense and Rep on the complementary sense. Genomoviridae is a recently established family of diverse circular ssDNA viruses. ![]() (Seoul, Korea) by primer walking and these sequence contigs were assembled using Geneious 11.1. The recombinant plasmids were Sanger sequenced at Macrogen Inc. The amplicons were resolved on a 0.7% agarose gel, amplicons of ~1.5–3 kb were excised, and gel purified, ligated in pJET1.2 vector (ThermoFisher Scientific, Waltham, MA, USA) and transformed into Escherichia coli XL1-Blue competent cells. For each PCR amplification, 1 µL of RCA product from each pooled sample was used as a template with primer pairs and Kapa HiFi Hotstart Ready Mix (Kapa Biosystems, Wilmington, MA, USA) using the following thermal cycling conditions: initial denaturation at 95 ☌ for 3 min, followed by 25 cycles at 98 ☌ for 20 s, 60 ☌ for 15 s and 72 ☌ for 3 min, final elongation at 72 ☌ for 3 min and a final renaturation at 4 ☌ for 10 min. These were used for PCR amplification of full genomes. Illumina Sequencing and Data ProcessingĪbutting primers were designed ( Supplementary Table S1) based on the de novo assembled contigs for genomoviruses and unclassified CRESS DNA. Viral DNA was extracted from 200 µL of resuspension using the Roche High Pure Viral nucleic acid kit (Roche Diagnostics, Indianapolis, IN, USA). The mixture was centrifuged at 15,000× g for 20 min and the resulting pellet was resuspended in 1 mL of filtrate. Louis, MO, USA) was added to 15 mL of each of the filtrates, the solution was mixed gently to suspend the PEG and incubated overnight at 4 ☌ to precipitate virions. The supernatant was filtered sequentially through a 0.45 µm, and 0.22 µm syringe filter. Briefly, approximately 5 g of fecal material per sample was homogenized in 20 mL SM buffer (0.1 M NaCl, 50 mM Tris-HCl, 10 mM MgSO 4) and centrifuged for 4700× g for 15 min. DNA was extracted as previously described by Steel et al. Samples were collected in the field from five locations in Maricopa, Yavapai, Pinal, and La Paz Counties, stored in separate plastic bags, and frozen once returned to the field station. Sonoran Desert tortoise fecal samples ( n = 40) were collected in the Sonoran Desert of Arizona, USA, in 20. Differences in the timing and amount of rainfall between the Mojave and Sonoran deserts may also have led to differential adaptation between these species over the same time period. They are thought to have speciated in isolation from Mojave desert tortoise ( Gopherus agassizii) approximately 5 MYA when the Colorado River bisected the ancestral population and began flowing into the Gulf of California. The Sonoran Desert tortoises eat a wide range of native desert grasses, are active in the summer monsoon, occupy rocky hillsides and streambeds laden with caliche caves, and usually use existing rock or caliche shelters. Spending much of their time underground in burrows or shelters, including months spent brumating during winter, they interact closely with desert soils and have commensal relationships with many desert animals (e.g., ground squirrels, wood rats, snakes, and spiders) through shared use of shelters. The Sonoran Desert tortoises ( Gopherus morafkai) are long-lived animals (>50 years in the wild) adapted to the Mojave and Sonoran deserts of southwestern North America. ![]()
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