Protocol: a simple gel-free method for SNP genotyping using allele-specific primers in rice and other plant species

<p>Abstract</p> <p>Background</p> <p>Genotype analysis using multiple single nucleotide polymorphisms (SNPs) is a useful but labor-intensive or high-cost procedure in plant research. Here we describe an alternative genotyping method that is suited to multi-sample or multi-locus SNP genotyping and does not require electrophoresis or specialized equipment.</p> <p>Results</p> <p>We have developed a simple method for multi-sample or multi-locus SNP genotyping using allele-specific primers (ASP). More specifically, we (1) improved the design of allele-specific primers, (2) established a method to detect PCR products optically without electrophoresis, and (3) standardized PCR conditions for parallel genomic assay using various allele-specific primers. As an illustration of multi-sample SNP genotyping using ASP, we mapped the locus for lodging resistance in a typhoon (<it>lrt5</it>). Additionally, we successfully tested multi-locus ASP-PCR analysis using 96 SNPs located throughout the genomes of rice (<it>Oryza sativa</it>) cultivars 'Koshihikari' and 'Kasalath', and demonstrated its applicability to other diverse cultivars/subspecies, including wild rice (<it>O. rufipogon</it>).</p> <p>Conclusion</p> <p>Our ASP methodology allows characterization of SNPs genotypes without electrophoresis, expensive probes or specialized equipment, and is highly versatile due to the flexibility in the design of primers. The method could be established easily in any molecular biology laboratory, and is applicable to diverse organisms.</p

Massive Intravascular Hemolysis in a Patient Infected by a Clostridium perfringens

Clostridium perfringens infection is a very rare cause of massive intravascular hemolysis, but it should always be kept in mind, since only early treatment can rescue patients from an otherwise rapidly fatal outcome. We report a case of a 78-year-old diabetic male who was admitted complaining of general fatigue, dark red urine, and vomiting. His blood revealed massive hemolysis. Computer tomography demonstrated huge liver abscess in the right lobe of the liver. About 1 h after admission, he suddenly fell into a critical condition. He died 3 h after admission in spite of intensive care and resuscitation. Clostridium perfringens was detected from the blood taken before death and from liver abscess by biopsy after death. We concluded that this patient died of acute massive intravascular hemolysis in septicemia caused by Clostridium perfringens infection

Retinal arterial macroaneurysm rupture caused by dissection-like change in the vessel wall

[Purpose] To highlight a potential pathogenetic mechanism of retinal arterial macroaneurysm. [Observations] A 79-year-old woman presented with a ruptured retinal arterial macroaneurysm in the right eye. One year after treatment, high-resolution multimodal imaging with optical coherence tomography (OCT), OCT angiography, and adaptive optics scanning light ophthalmoscope showed that a narrow passage developed in the vessel wall of the pre-existing retinal arterial macroaneurysm and another macroaneurysm had developed adjacent to the pre-existing one. [Conclusions and Importance] These images suggest the pathogenesis of retinal arterial macroaneurysm (RAM) associated with crack-like changes in the retinal arterial wall, similar to pathologies seen in systemic arteries

Western North Pacific tropical cyclone model tracks in present and future climates

Western North Pacific tropical cyclone (TC) model tracks are analyzed in two large multimodel ensembles, spanning a large variety of models and multiple future climate scenarios. Two methodologies are used to synthesize the properties of TC tracks in this large data set: cluster analysis and mass moment ellipses. First, the models' TC tracks are compared to observed TC tracks' characteristics, and a subset of the models is chosen for analysis, based on the tracks' similarity to observations and sample size. Potential changes in track types in a warming climate are identified by comparing the kernel smoothed probability distributions of various track variables in historical and future scenarios using a Kolmogorov-Smirnov significance test. Two track changes are identified. The first is a statistically significant increase in the north-south expansion, which can also be viewed as a poleward shift, as TC tracks are prevented from expanding equatorward due to the weak Coriolis force near the equator. The second change is an eastward shift in the storm tracks that occur near the central Pacific in one of the multimodel ensembles, indicating a possible increase in the occurrence of storms near Hawaii in a warming climate. The dependence of the results on which model and future scenario are considered emphasizes the necessity of including multiple models and scenarios when considering future changes in TC characteristics

DRUG-ADMINISTERING PERSONS' EXPOSURE TO ORAL ANTICANCER DRUGS TO BE ADMINISTERED THROUGH A TUBE

ABSTRACTObjective: The objective of this study was to quantitatively evaluate anticancer drug exposure of non-health care professionals who administer drugsthrough a tube employing a method devised by us.Methods: The subjects were 30 general volunteers aged 22-84 years. They wore gloves and administered Indian ink, simulating an anticancer drug, toa multipurpose adult human-type patient care simulator through a tube using 5 types of syringe, and the area stained with Indian ink was measured.Results: When comparing the number of pixels among the syringes regardless of age, Syringe B showed the lowest number (11.8±3.1 cm2), and therewas a significant difference between Syringes B and E. Furthermore, we compared the total number of pixels in each age group regardless of the typeof syringe. In the 20-year-old group, it was the lowest (10.9±2.3 cm2) showing significant differences in comparison with the other groups. WhenSyringe B was used, the number of pixels was markedly lower than on adopting the other syringes.Conclusion: It was clarified that the level of exposure to anticancer drugs markedly varies depending on the type of syringe and age. It was alsoclarified that the method to evaluate exposure to anticancer drugs using Indian ink devised by us is simple and useful.Keywords: Oral anticancer drugs, Simple suspension method, Drug-administering persons' exposure

DETECTION AND CHARACTERISTICS OF UNRUPTURED RETINAL ARTERIAL MACROANEURYSMS

Purpose: To determine the presence of unruptured retinal arterial macroaneurysms (RAMs) and to examine the characteristics of the detected lesions. Methods: This retrospective observational study included the affected and contralateral eyes of 50 patients (100 eyes) with symptomatic, unilateral, ruptured RAMs who visited the Department of Ophthalmology at the Kyoto University Hospital (April 2014–April 2020) and were followed up for at least 6 months after the onset. The presence and characteristics of unruptured RAMs were examined by reviewing the findings of color fundus photography and infrared scanning laser ophthalmoscopy performed before the onset or during the follow-up period. Results: Unruptured RAMs were detected in six of the 50 patients. Some patients had bilateral or multiple unruptured RAMs, and a total of 12 unruptured RAMs were detected in eight eyes of the six patients. Among the detected lesions, eight exhibited a longitudinal increase in their diameter during the follow-up period, whereas six exhibited ruptures. Conclusion: Unruptured RAM is not an uncommon retinal vascular abnormality and can enlarge and progress to ruptured RAM

DRUG-ADMINISTERING PERSONS' EXPOSURE TO ORAL ANTICANCER DRUGS TO BE ADMINISTERED THROUGH A TUBE

Objective: The objective of this study was to quantitatively evaluate anticancer drug exposure of non-health care professionals who administer drugs through a tube employing a method devised by us. Methods: The subjects were 30 general volunteers aged 22-84 years. They wore gloves and administered Indian ink, simulating an anticancer drug, to a multipurpose adult human-type patient care simulator through a tube using 5 types of syringe, and the area stained with Indian ink was measured. Results: When comparing the number of pixels among the syringes regardless of age, Syringe B showed the lowest number (11.8±3.1 cm2), and there was a significant difference between Syringes B and E. Furthermore, we compared the total number of pixels in each age group regardless of the type of syringe. In the 20-year-old group, it was the lowest (10.9±2.3 cm2) showing significant differences in comparison with the other groups. When Syringe B was used, the number of pixels was markedly lower than on adopting the other syringes. Conclusion: It was clarified that the level of exposure to anticancer drugs markedly varies depending on the type of syringe and age. It was also clarified that the method to evaluate exposure to anticancer drugs using Indian ink devised by us is simple and useful

MACULAR BLOOD FLOW CHANGES IN BRANCH RETINAL VEIN OCCLUSION EXAMINED BY OPTICAL COHERENCE TOMOGRAPHY ANGIOGRAPHY VARIABLE INTERSCAN TIME ANALYSIS

Purpose: To examine the relationship between changes in retinal blood flow and the recurrence of macular edema in eyes with branch retinal vein occlusion. Methods: This observational study included 32 eyes in 32 patients (18 men and 14 women) with branch retinal vein occlusion who visited the Department of Ophthalmology at Kyoto University Hospital (February 2021–November 2021). At the time of inclusion in the study, each patient underwent optical coherence tomography angiography on a macular area measuring 4 × 4 mm2. For variable interscan time analysis, different interscan times were set at 7.6 (IST7.6) and 20.6 ms (IST20.6) for the optical coherence tomography angiography. The parafoveal vessel densities were measured sectorally at IST7.6 and IST20.6, and their relationship with the longitudinal changes evident in the retinal thicknesses during the variable interscan time analysis examination and 2 months later was evaluated. Results: The parafoveal vessel densities in the affected sector was significantly greater at IST20.6 than at IST7.6 (P = 0.011). At 2 months after the variable interscan time analysis examination, 6 patients (19%) showed recurrence of macular edema involving the fovea. The difference in the parafoveal vessel densities (IST20.6 − IST7.6) in the affected sector was significantly associated with longitudinal retinal thickening in the corresponding parafovea (P = 0.020) and fovea (P = 0.014). Conclusion: In eyes with branch retinal vein occlusion, optical coherence tomography angiography variable interscan time analysis facilitated the detection of retinal blood flow changes that might be predictive for the recurrence of macular edema

Foveal Thickness Fluctuation in Anti-VEGF Treatment for Branch Retinal Vein Occlusion: A Long-term Study

PURPOSE: Branch retinal vein occlusion (BRVO) causes macular edema (ME), which can be controlled with anti-VEGF treatments. However, these treatments are not curative, necessitating additional anti-VEGF treatments at recurrence. Long-term results, optimal anti-VEGF treatment regimens, and the comprehensive effects of ME recurrence are largely unknown. Thus, we aimed to examine the effects of foveal thickness (FT) fluctuation (FTF) on the visual and morphologic outcomes of anti-VEGF treatments for BRVO-ME administered via a pro re nata regimen. DESIGN: A retrospective, observational case series. SUBJECTS: This study analyzed 309 treatment-naïve patients (309 eyes) with BRVO-ME between 2012 and 2021 at a multicenter retinal practice. METHODS: The FT was assessed using OCT at each study visit. MAIN OUTCOME MEASURES: We evaluated the logarithm of the minimal angle of resolution (logMAR) best corrected visual acuity (BCVA) and the defect length of the foveal ellipsoid zone (EZ) band using OCT. RESULTS: At baseline, the mean logMAR BCVA was 0.30 ± 0.30 and the mean FT was 503 ± 162 μm. The number of anti-VEGF injections for BRVO-ME was 5.8 ± 4.6 during the mean follow-up period (50.6 ± 22.2 months). At the final examination, the mean logMAR BCVA and FT values were significantly improved compared with those at the baseline. Multiple regression analyses showed that age, baseline logMAR BCVA, and FTF were significantly associated with the final logMAR BCVA (β = 0.20, 0.35, and 0.30, respectively). Foveal thickness fluctuation (divided into groups 0-3 in ascending order of FTF) was significantly associated with logMAR BCVA and the defect length of the foveal EZ band at the final examination. The defect lengths of the foveal EZ band were longitudinally shortened in groups 0 and 1 and were slightly prolonged in groups 2 and 3. The logMAR BCVA showed improvements in groups 0 and 1 and worsened slightly in groups 2 and 3. CONCLUSIONS: Foveal thickness fluctuation was significantly associated with visual acuity and foveal photoreceptor status. Thus, the morphologic and functional prognoses of eyes with BRVO may improve with the identification of the characteristics of eyes with greater FTF and consequently controlling the FTF more strictly

Giantin Affects Golgi Stack Connection

Golgins are a family of Golgi-localized long coiled-coil proteins. The major golgin function is thought to be the tethering of vesicles, membranes, and cytoskeletal elements to the Golgi. We previously showed that knockdown of one of the longest golgins, Giantin, altered the glycosylation patterns of cell surfaces and the kinetics of cargo transport, suggesting that Giantin maintains correct glycosylation through slowing down transport within the Golgi. Giantin knockdown also altered the sizes and numbers of mini Golgi stacks generated by microtubule de-polymerization, suggesting that it maintains the independence of individual Golgi stacks. Therefore, it is presumed that Golgi stacks lose their independence following Giantin knockdown, allowing easier and possibly increased transport among stacks and abnormal glycosylation. To gain structural insights into the independence of Golgi stacks, we herein performed electron tomography and 3D modeling of Golgi stacks in Giantin knockdown cells. Compared with control cells, Giantin-knockdown cells had fewer and smaller fenestrae within each cisterna. This was supported by data showing that the diffusion rate of Golgi membrane proteins is faster in Giantin-knockdown Golgi, indicating that Giantin knockdown structurally and functionally increases connectivity among Golgi cisternae and stacks. This increased connectivity suggests that contrary to the cis-golgin tether model, Giantin instead inhibits the tether and fusion of nearby Golgi cisternae and stacks, resulting in transport difficulties between stacks that may enable the correct glycosylation of proteins and lipids passing through the Golgi