Detection of the genetic variation of polygalacturonase-inhibiting protein gene 2 in autotetraploid alfalfa (Medicago sativa) using an improved SSCP technique.

In this study, 2 approaches were adopted to obtain good single-strand conformation polymorphism (SSCP) data for autotetraploid alfalfa; primers were added to PCR products, and fluorescent-labeled primers were utilized. PCR-SSCP conditions for a 331-bp fragment in the coding region of polygalacturonase-inhibiting protein gene 2 in alfalfa (MsPGIP2) were optimized, and the results showed that the best SSCP gel pattern could be obtained when the loading mixture was made by mixing 1 µL PCR products, 0.2 to 0.8 µL unlabeled primers (50 µM) and 4 to 16 µL loading buffer. Furthermore, the use of the fluorescent-labeled primers resulted in 2 separated electrophoresis images from 2 complementary single DNA strands, thus making the determination of alleles and idiotypes a relatively easy task. In addition, the results of sequencing prove that the determination of alleles and idiotypes were accurate based on SSCP analysis. Finally, a total of 9 alleles with 18 SNP sites were identified for MsPGIP2 in the alfalfa variety 'Algonquin'. In conclusion, MsPGIP2 possessed great genetic variation, and the addition of primers to the PCR products in combination with the fluorescent labeling of primers could significantly improve the sensitivity and resolution of SSCP analysis. This technique could be used for genetic diversity detection and marker-assisted breeding of useful genes in autopolyploid species such as alfalfa.

Genome-wide multilocus analysis of intraspecific differentiation in Oryza rufipogon Griff. from China and the influence of introgression from O. sativa L.

Twenty-five populations of Oryza rufipogon from China and 144 cultivars of Oryza sativa were selected for this study. Based on the DNA fragment of Ehd1-4 and subspecies-specific sequence-tagged site markers in different chromosomes, intraspecific differentiation in O. rufipogon from China was analyzed. The introgression from O. sativa to O. rufipogon was also analyzed based on simple sequence repeat markers. The results revealed that the DNA fragment of Ehd1-4 could distinguish the O. sativa subspecies japonica and indica. Furthermore, although significant indica-japonica differentiation did not occur in most O. rufipogon populations from China, O. rufipogon varieties from Hainan Island and from the mainland of China showed differentiation tendencies. Japonica-like O. rufipogon varieties were predominant in Mainland China. However, more indica-like O. rufipogon varieties were found in Hainan Island. Finally, although cultivar-specific alleles were found in most of the O. rufipogon varieties from Hainan Island and Guangdong Province, some varieties remain pure and non-introgressive.

Association of AFLP and SCAR markers with common leafspot resistance in autotetraploid alfalfa (Medicago sativa).

To identify amplified fragment length polymorphism (AFLP) markers associated with resistance or susceptibility of alfalfa to common leafspot (CLS) caused by the fungus Pseudopeziza medicaginis (Dermateaceae), bulked segregant analysis was conducted based on an F(1(M × M)) population of 93 plants and a BC(1)S population of 91 plants. Three AFLP markers, ACTCAA(R206), TAGCAC(R185), and GGACTA(S264), were found to be associated with CLS resistance or susceptibility. All three markers were found at significantly different frequencies (71.9, 80.3 and 91.8%) compared to resistant or susceptible plants in the original population. Subsequently, these three AFLP markers were converted into three SCAR markers, ACTCAA(R136), TAGCAC(R128) and GGACTA(S254), which are easier to employ in breeding programs. The three SCAR markers were used in a randomly selected population with 50% resistance; the probability of finding one resistant plant was increased to 67.3, 66.7 and 90.0% with markers ACTCAA(R136), TAGCAC(R128) and GGACTA(S254), independently. If two of the SCAR markers were used simultaneously, the probability would be higher than 89%. The three SCAR markers identified in this study would be applicable for selection for CLS resistance in alfalfa breeding programs. Moreover, the genetic analysis indicated that CLS resistance in alfalfa is conferred by a single dominant gene.

Dynamic factors in the reactions between the magic cluster Al(13)⁻ and HCl/HI.

The icosahedral Al is a "magic" cluster with remarkable stability due to its high symmetry and closed valence shells. Its reactivity has provided a molecular model for understanding oxidation and dissolution processes in bulk metals. By first principles calculations, we demonstrated the importance of dynamic factors in the Al + HX reactions, with HX being either HCl or HI. There was a barrier to the dissociative adsorption of HX on the surface of an Al cluster, which involved charge transfer from Al. Furthermore, the H atom could be bonded to the cluster in multiple ways, similar to the top, bridge and hollow adsorption sites on Al(111) surface. With a large amount of energy (∼40 kcal mol(-1)) deposited during the formation of Al(13)HX(-), the H atom could easily migrate among these sites, similar to the diffusion of hydrogen on metal surfaces. These factors were therefore important considerations in the formation and dissociation of Al(13)HX(-), and more generally in reactions involving other metal clusters.

[Evaluation of Schneiderian membrane state using fiber optic endoscope during maxillary sinus floor elevation with lateral window].

Objective: To observe the status of the sinus membrane using fiber optic endoscope during the lateral window approach sinus floor elevation to provide a reference for clinicians when evelvating the sinus mucoperiosteum. Methods: Sixty-six patients (72 sides) who underwent maxillary sinus floor elevation in Beijing Ruicheng Stomatology Hospital from September 2016 to December 2019 were selected, including 40 males and 26 females, aged 26-80 years old [(56.2±11.5) years]. And fiber optic endoscopy was used to observe the maxillary mucoperiosteum during the operation. Results: The status of maxillary sinus mucoperiosteal during lateral window approach sinus floor elevation can be divided into four categories: ① Class Ⅰ, complete periosteal, no damage to mucoperiosteum; ②Class Ⅱ, periosteal injury, unexposed laminae propria; ③Class Ⅲ, periosteal Rupture, exposed lamina propria; ④ Class Ⅳ, mucoperiosteum perforation, rupture of periosteum, lamina propria and epithelial layer. A total of 72 operations were performed, including 18 cases of class I, 28 cases of class Ⅱ, 4 cases of class Ⅲ, and 22 cases of class Ⅳ. Conclusions: The status of maxillary sinus mucoperiosteal during lateral window approach sinus floor elevation can be divided into four categories. Fiberoptic endoscopy as a clinical auxiliary examination method can improve the operator's control of the status of the maxillary sinus membrane and assist the peeling of the mucosa.

Cannabinoid receptor 2 activation alleviates septic lung injury by promoting autophagy via inhibition of inflammatory mediator release.

Septic lung injury is one of main causes of high mortality in severe patients. Inhibition of excessive inflammatory response is considered as an effective strategy for septic lung injury. Previous studies have shown that cannabinoid receptor 2 (CB2), a G protein-coupled receptor, play an important role in immunosuppression. Whether CB2 can be used as a therapeutic target for septic lung injury is unclear. The aim of this study is to explore the role of CB2 in sepsis and its potential mechanism. In this study, treatment with HU308, a specific agonist of CB2, could reduce lung pathological injury, decrease the level of inflammatory cytokines and strengthen the expression of autophagy-related gene after cecal ligation puncture (CLP)-induced sepsis in mice. Similar results were obtained in RAW264.7 macrophages after LPS treatment. Furthermore, the effect of HU308 could be blocked by autophagy blocker 3-MA in vivo and in vitro. These results suggest that CB2 serves as a protective target for septic lung injury by decreasing inflammatory factors, which is associated with the enhancement of autophagy.

Histamine H3 receptor antagonist Clobenpropit protects propofol-induced apoptosis of hippocampal neurons through PI3K/AKT pathway.

OBJECTIVE: The aim of the study was to explore whether histamine H3 receptor antagonist Clobenpropit could protect propofol-induced neurotoxicity in hippocampal neurons. MATERIALS AND METHODS: Hippocampal neurons were extracted from neonatal rats and induced with propofol. MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide) assay was performed to detect apoptotic rate of neurons. Western blot was conducted to detect protein levels of cleaved-caspase-3 and Bax/Bcl2. After LY294002 treatment, the PI3K pathway antagonist was applied in neurons, protein levels of cleaved-caspase-3 and Bax/Bcl2 were detected by Western blot as well. RESULTS: Propofol treatment remarkably induced neuronal apoptosis. Clobenpropit alleviated cell apoptosis induced by propofol. Protein expressions of cleaved-caspase-3 and Bax/Bcl2 were remarkably downregulated in neurons treated with Clobenpropit. LY294002 induction remarkably reverses the protective role of Clobenpropit in neuronal apoptosis, manifesting as downregulated PI3K and p-AKT after LY294002 treatment. CONCLUSIONS: Clobenpropit protects propofol-induced neuronal apoptosis through activating PI3K/AKT pathway.

L-Cysteine promotes the proliferation and differentiation of neural stem cells via the CBS/H2S pathway.

Growing evidence has suggested that hydrogen sulfide (H2S) acts as a novel neuro-modulator and neuroprotective agent; however, it remains to be investigated whether H2S has a direct effect on neural stem cells (NSCs). We report here that NSCs expressed cystathionine ß synthase (CBS) and addition of exogenous H2S donor, L-cysteine, stimulated proliferation and increased the differentiation potential of NSCs to neurons and astroglia. Moreover, pre-treatment with aminooxyacetic acid, the inhibitor of CBS or knockdown of CBS in using siRNA, significantly attenuated the effects of L-cysteine on elevated H2S levels and the cell proliferation; it also effectively suppressed L-cysteine-induced neurogenesis and astrocytogenesis. Further analysis revealed that L-cysteine-induced proliferation was associated with phosphorylation of extracellular signal-regulated kinases 1/2 and differentiation with altered expression of differentiation-related genes. Taken together, the present data suggest that L-cysteine can enhance proliferation and differentiation of NSCs via the CBS/H2S pathway, which may serve as a useful inference for elucidating its role in regulating the fate of NSCs in physiological and pathological settings.

Investigation of rice transgene flow in compass sectors by using male sterile line as a pollen detector.

Rice is the most important staple food in the world. The rapid development of transgenic rice and its future commercialization have raised concerns regarding transgene flow and its potential environmental risk. It is known that rice is a self-pollinated crop; the outcrossing rate between common cultivars is generally less than 1%. In order to improve the detection sensitivity of rice transgene flow, a male sterile (ms) line BoA with a high outcrossing rate was used as a pollen detector in this study. A concentric circle design was adopted, in which the transgenic rice B2 containing bar gene as a pollen donor was planted in the center circle and the recipient BoA was planted in eight compass sectors. The frequency of transgene flow in compass sectors was analyzed by continuous sampling to generate cumulative data. The results of two years with sound reproducibility demonstrated that the rice gene flow was closely associated with the wind direction. According to the mean frequency of transgene flow, the eight sectors can be divided into two groups: a higher frequency group downstream of the prevailing wind (DPW) with a mean frequency ranging from 6.47 to 26.24%, and a lower frequency group lateral to or upstream of the prevailing wind (UPW) with a mean frequency of 0.39 to 3.03%. On the basis of the cumulative data, 90-96% of the cumulative gene flow events occurred in the four DPW sectors, while it was 4-10% in the four UPW sectors. By using these systematic data, simulation models and isograms of transgene flow in the eight compass sectors were calculated and drawn, respectively.