A mutation-sensitive, multiplexed and amplification-free detection of nucleic acids by stretching single-molecule tandem hairpin probes.

The detection of nucleic acid sequences in parallel with the discrimination of single nucleotide variations (SNVs) is critical for research and clinical applications. A few limitations make the detection technically challenging, such as too small variation in probe-hybridization energy caused by SNVs, the non-specific amplification of false nucleic acid fragments and the few options of dyes limited by spectral overlaps. To circumvent these limitations, we developed a single-molecule nucleic acid detection assay without amplification or fluorescence termed THREF (hybridization-induced tandem DNA hairpin refolding failure) based on multiplexed magnetic tweezers. THREF can detect DNA and RNA sequences at femtomolar concentrations within 30 min, monitor multiple probes in parallel, quantify the expression level of miR-122 in patient tissues, discriminate SNVs including the hard-to-detect G-U or T-G wobble mutations and reuse the probes to save the cost. In our demonstrative detections using mock clinic samples, we profiled the let-7 family microRNAs in serum and genotyped SARS-CoV-2 strains in saliva. Overall, the THREF assay can discriminate SNVs with the advantages of high sensitivity, ultra-specificity, multiplexing, reusability, sample hands-free and robustness.

[Temporal variability of the isotope compositions of plum rain in Nanjing from event to interannual time scales]. / å—äº¬æ¢ é›¨é™æ°´åŒä½ç´ ç»„æˆä»Žäº‹ä»¶åˆ°å¹´é™ å°ºåº¦çš„å˜åŒ–ç‰¹å¾.

Precipitation in the plum rain period accounts for 40%-50% of annual precipitation in the monsoon region. To clarify the temporal variability of the isotopic composition of precipitation during the plum rain period from event to interannual time scale and identify the influencing factors, we analyzed the isotopic composition of precipitation and its influencing factors in Nanjing from 2015 to 2022. By using the Hybrid Single-particle Lagran-gian Integrated Trajectory (HYSPLIT) model with specific humidity analysis, we investigated the water vapor source and influencing factors. The results showed that 1) the isotopic abundance of atmospheric precipitation was depleted in the summer and enriched in winter. dx was lower in summer and higher in winter. The isotopic abundance of precipitation from the plum rain was depleted compared to mean value of the whole-year. 2) There was no significant correlation between δ2H and δ18O of the plum rain (precipitation) with local meteorological factors. However, dx was lower in light rain, reflecting the effect of sub-cloud evaporation. The average dx was higher during plum rain period in years with more total plum rain precipitation. 3) The low-latitude South China Sea and the western Pacific Ocean source area provided water vapor for the plum rain. The shift of moisture source region led to abrupt changes in precipitation isotopes. Our results could provide data support for studies on precipitation isotopes in the monsoon region, as well as a reference point for further understanding the precipitation mechanism of the plum rain and stu-dying the seasonal variability of atmospheric circulation in the East Asian monsoon region.

[Effects of Water Vapor Source and Local Evaporation on the Stable Hydrogen and Oxygen Isotopic Compositions of Precipitation].

Stable hydrogen and oxygen isotopic compositions in precipitation are good tracers and can provide unique information about the water cycle. Precipitation samples were collected at the Nanjing, Liyang, Yixing, and Dongshan sites in 2016, and the HDO and H218O compositions of precipitation were measured. The temporal variability of HDO and H218O compositions and deuterium-excess of precipitation were analyzed, and the influence of the water vapor source and local evaporation on stable isotopic composition of precipitation were discussed. The results indicated that:① Seasonal variations in the HDO composition, H218O composition, and deuterium-excess of precipitation occurred due to different water vapor sources during the summer and winter monsoon seasons. The HDO and H218O compositions were depleted during the summer monsoon season and enriched during the winter monsoon season. The deuterium-excess during the summer monsoon season was lower compared to the winter monsoon season. ② During the summer monsoon, the evaporation of Lake Taihu made the deuterium-excess of downwind precipitation and the downwind intercept of the local meteoric water line higher. During the winter monsoon season, local evaporation had little influence on HDO and H218O components in precipitation. ③ Both of the intercepts and slopes of the local meteoric water line were higher than those of the global meteoric water line, due to moisture recycling during the winter monsoon season and different water vapor sources between the summer and winter monsoon seasons.

[Quantification of Methane Ebullition Flux from Small Ponds Using the Inverted-Funnel Method].

To quantify the ratio of CH4 ebullition to total flux in subtropical shallow ponds, the CH4 flux at the water-air interface was measured using the inverted-funnel and water equilibrium methods in two small ponds in Quanjiao, Anhui Province from July 28 to August 13, 2016. The average CH4 ebullition fluxes were 121.78 and 161.08 mg·(m2·d)-1 and the average diffusion fluxes were 3.38 and 3.79 mg·(m2·d)-1 over pond A and pond B, respectively. The ebullition flux accounted for 97.5% and 96.4% of the total flux over pond A and pond B, respectively. Methane ebullition ranged from 0.11 to 446.90 mg·(m2·d)-1 over pond A and from 0.05 to 607.51 mg·(m2·d)-1 over pond B. Gas ebullition rate during the day was higher than that at night and was controlled by wind speed. Methane ebullition flux was influenced by wind speed over the shallow pond at hourly scale and by water depth and wind speed at daily scale, with positive correlation with wind speed and negative correlation with water depth. Varying with latitude, methane ebullition flux was higher for the water bodies in the mid-latitude region compared to those in the high-latitude region. Direct observations of the methane ebullition flux over small ponds provide data support and theoretical reference to precisely estimate the contribution of inland water bodies to regional and global carbon cycle.

Boron deficiency decreases growth and photosynthesis, and increases starch and hexoses in leaves of citrus seedlings.

Seedlings of sweet orange (Citrus sinensis) were fertilized for 14 weeks with boron (B)-free or B-sufficient (2.5 or 10 microM H(3)BO(3)) nutrient solution every other day. Boron deficiency resulted in an overall inhibition of plant growth, with a reduction in root, stem and leaf dry weight (DW). Boron-starved leaves showed decreased CO(2) assimilation and stomatal conductance, but increased intercellular CO(2) concentrations. Activities of ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco), NADP-glyceraldehyde-3-phosphate dehydrogenase (NADP-GAPDH) and stromal fructose-1,6-bisphosphatase (FBPase) were lower in B-deficient leaves than in controls. Contents of glucose, fructose and starch were increased in B-deficient leaves while sucrose was decreased. Boron-deficient leaves displayed higher or similar superoxide dismutase (SOD), ascorbate peroxidase (APX), monodehydroascorbate reductase (MDAR) and glutathione reductase (GR) activities, while dehydroascorbate reductase (DHAR) and catalase (CAT) activities were lower. Expressed on a leaf area or protein basis, B-deficient leaves showed a higher ascorbate (AsA) concentration, but a similar AsA concentration on a DW basis. For reduced glutathione (GSH), we found a similar GSH concentration on a leaf area or protein basis and an even lower content on a DW basis. Superoxide anion (O(2)(-)) generation, malondialdehyde (MDA) concentration and electrolyte leakage were higher in B-deficient than in control leaves. In conclusion, CO(2) assimilation may be feedback-regulated by the excessive accumulation of starch and hexoses in B-deficient leaves via direct interference with chloroplast function and/or indirect repression of photosynthetic enzymes. Although B-deficient leaves remain high in activity of antioxidant enzymes, their antioxidant system as a whole does not provide sufficient protection from oxidative damage.

[Pathomorphological observation of the hypertrophic scar induced by injury to conical structure in female red Duroc pig].

UNLABELLED: To further explore the relationship between hypertrophic scar and injury to conical structure of skin and the pathogenesis of hypertrophic scar, and to reproduce an optimal animal model of hypertrophic scar. METHODS: The back of two FRDP pigs were shaved, and a piece of normal skin was harvested for the observation of conical structure of skin. Skin wounds with depth of 0.38 mm, 0.76 mm, 1.14 mm and 1.52 mm, respectively, were created by gas-driven dermatome. Eight wounds measuring 7.0 cm x 7.0 cm were created on each pig. The wounds were divided into 4 groups according to the wound depth with 4 wounds in each group, i.e. 0.38 mm group, 0.76 mm group, 1.14 mm group and 1.52 mm group. The 0.38 mm and 0.76 mm groups were designated as superficial wound groups and 1.14 mm and 1.52 mm groups as deep wound groups. The wounds were allowed to heal without treatment. Tissue samples from the wound were harvested on 0, 10, 30, 60, 90 and 150 post injury day (PID) , and they were sectioned for HE staining and staining for elastic fibers (VVG). The wound healing and the scar formation were observed with naked eye. The skin conical structures in normal and injured skin were also observed. The morphology of hypertrophic scar was observed, and the thickness of the scar tissue was determined and scored. RESULTS: The wounds in superficial wound groups healed within 3 weeks with flat surface without scar formation. The wounds in deep wound groups healed later than 4 weeks with thick, hairless, hard in texture, with depigmentation or pigmentation, finally forming contracture. The skin conical structure could be found on the back of FRDP with HE and VVG staining, and it was similar to that of human in terms of the structure. In superficial wound groups, the upper part of the skin conical structure was injured, but fat fornix and glands were intact. In deep wound groups, the lower part of the skin cone, together with the fat fornix and gland were all injured. On the 150th post injury day, the histological picture of the tissue in superficial wound groups was similar to that of normal skin. But the skin conical structure could not be found in deep wound groups, and the wounds were filled by a large accumulation of disarrayed and irregularly arranged collagen fibers. With passage of time, the scar became thicker and thicker, and the scar hypertrophy reached the zenith in 150th PID. CONCLUSION: The injury of skin conical structure can lead to the formation of hypertrophic scar. FRDP can be used to reproduce and ideal model of hypertrophic scar.