多个抗虫基因转化水稻两用系培矮—64S

应用基因枪法对水稻两用系培矮－64S进行了转化。质粒载体pKC-3串联了三个抗虫基因,将其转化成熟胚诱导形成的愈伤组织,共获得33株转基因植株。分别对R0代植株不同的基因进行PCR及Southern blot分析,并对R1代植株进行了PCR分析。结果表明,三个抗虫基因均已整合到水稻基因组并获得稳定遗传。     

植物性雌激素genistein对豚鼠乳头肌的电生理效应

应用细胞内微电极技术,观察了genistein(GST)对豚鼠乳头肌的电生理效应。结果显示：（1）GST（10-100μmol/L)浓度依赖地缩短正常乳头肌动作电位时程;（2）对部分去极化乳头肌,GST（50μmol/L)除缩短动作电位时程外,还使动作电位幅值和超射值降低,零相最大上升速度减慢;（3）预先应用一氧化氮合酶抑制剂L-NNA（5mmol/L)。不影响GST（50μmol/L)的电生理效应;（4）单独应用17β-雌二醇（E2,5μmol/L)或GST（10μmol/L)时,动作电位各参数无明显变化。而预先应用同剂量的GST再加入E2,则动作电位时程缩短,结果提示,GST可能通过非NO途径抑制Ca^2 内流,从而影响豚鼠乳头肌电生理效应,并与E2有加强或协同效应。     

侵染中华蜜蜂6日龄幼虫的蜜蜂球囊菌的微小RNA差异表达谱及调控网络

【目的】蜜蜂球囊菌(Ascosphaeraapis,简称球囊菌)是专性侵染蜜蜂幼虫的致死性真菌病原。MicroRNA(miRNA)作为一类重要的基因表达调控因子,能够广泛参与真菌及其宿主的相互作用过程。本研究通过比较分析球囊菌孢子(AaCK)和侵染中华蜜蜂(Apis cerana cerana,简称中蜂) 6日龄幼虫肠道内的球囊菌(AaT)的smallRNA(sRNA)组学数据对球囊菌的差异表达miRNA(differentiallyexpressed miRNA,DEmiRNA)、靶mRNA及二者间的调控网络进行全面解析,旨在揭示miRNA介导的球囊菌对中蜂幼虫的侵染机制。【方法】对于球囊菌侵染的中蜂6日龄幼虫肠道的small RNA-seq (sRNA-seq)数据,利用BLAST工具连续比对东方蜜蜂(Apiscerana)和球囊菌的参考基因组筛滤得到AaT的sRNA组学数据。分别将AaCK和AaT的sRNA组学数据比对miRBase数据库,对球囊菌侵染宿主前后miRNA的数量和结构特征进行分析。联用RNAhybrid+svm_light、Miranda和Tar...     

Characteristics of soil water distribution and evaluation of recharge rate under different grazing history in the Xilin Gol Steppe北大核心CSCD

Aims In the Xilin Gol Steppe, human-induced grassland degradation and land desertification are becoming increasingly severe. Critical evaluation of its impact on soil water and recharge rate is important for sustainable management of soil health and water resources in the region. Methods In order to determine the effect of different grazing history on dynamics of deep soil moisture contents and precipitation infiltration in the Xilin Gol Steppe, three sites with different grazing history (ungrazed since 1979 or UG79, ungrazed since 1999 or UG99, and continuously grazed or CG) were selected with two sampling spots for each site. The precipitation infiltration was estimated using the chloride mass balance method. Important findings The results showed that: 1) Average soil water content of 0–5 m was 7.1%, 6.9%, and 6.3% for UG79, UG99, and CG, respectively, with no significant difference. In the soil layer of 0–2 m, the soil water content of UG79 was 26.6% and 33.7% higher than that of UG99 and CG, respectively. The soil water content of UG79 was significantly higher than that of UG79 and UG99 (p < 0.05) with no significant difference between UG99 and CG. The soil water storage capacity of UG79 was 87.19 mm higher than UG99 and 82.52 mm higher than CG. In the deep layer of 2–5 m, no significant difference in the soil water content and the water storage among different grazing history. 2) The factors influencing soil water differed among different grazing treatments. The soil water content was mainly affected by the vegetation conditions and soil properties for the 0–2 m soil layer, but by the composition of soil particles for the 2–5 m soil layer. The effect of soil organic matter (SOM) content on soil water increased with time without grazing. Soil water content of the entire soil profile of UG79 was significantly correlated with soil texture and SOM content (p < 0.01). Soil water content of 0–2 m was significantly correlated with SOM content (p < 0.01), soil water content of 2–5 m was significantly correlated with the soil texture (p <0.01), but soil moisture content of UG99 and CG had no significant correlation with SOM content. 3) Annual recharge rate was 5.64, 3.54, and 2.45 mm·a –1 for UG79, UG99 and CG, respectively. The recharge rate increased by 44.5% and 130.2% for the site without grazing for 15 and 35 years, respectively. The recharge rate in the study area ranged from 1.95 to 7.61 mm·a –1 , accounting for only 0.55%–2.13% of the precipitation. In summary, ungrazing treatment can increase soil water retention, total water storage capacity, and recharge. © 2018 Editorial Office of Chinese Journal of Plant Ecology. All Rights Reserved.     

c-di-GMP的磷酸二酯酶PA4781在抗菌肽Merecidin抑制铜绿假单胞菌生物被膜中的作用

【背景】抗菌肽Merecidin可抑制临床菌株铜绿假单胞菌PA03生物被膜。PA4781基因是课题组通过生物信息学分析筛选出的差异表达基因,PA4781作为细菌第二信使分子环二鸟苷酸(cyclic diguanylate,c-di-GMP)的磷酸二酯酶具有降解c-di-GMP的作用,其在抗菌肽Merecidin抑制生物被膜中的作用机制尚不清楚。【目的】研究细菌第二信使分子c-di-GMP的磷酸二酯酶PA4781基因在抗菌肽Merecidin抑制铜绿假单胞菌生物被膜中的作用。【方法】利用单碱基突变技术敲除PA4781基因,Sanger测序方法检测敲除的正确性。采用结晶紫染色法观察PA03菌株、PA4781过表达菌株、PA4781敲除菌株24 h生物被膜生长情况,以及在抗菌肽Merecidin 24、48、72μmol/L作用下各菌株生物被膜的生长情况。采用对羟基联苯溶液显色法检测在抗菌肽Merecidin 48、72μmol/L作用下,PA03菌株、PA4781过表达菌株、PA4781敲除菌株生物被膜藻酸盐的变化情况。【结果】Sanger测序结果显示,用pnCasPABEC系统成功实现了...     

Analysis on additive effects and additive-by-additive epistatic effects of QTLs for yield traits in a recombinant inbred line population of rice

A linkage map consisting of 158 DNA markers were constructed by using a recombinant inbred line (RIL) population derived from the indica-indica rice cross Zhenshan 97B 2 Milyang 46. Quantitative trait loci (QTLs) conditioning grain yield and five yield component traits were determined at the one-locus and two-locus levels, and genotype-by-environment (GE) interactions were analyzed. Thirty-one QTLs were detected to have significant additive effects for yield traits, of which 12 also exhibited significant epistatic effects. Sixteen significant additive-by-additive (AA) interactions were detected, of which nine occurred between QTLs with own additive effects (M_epQTLs), four occurred between QTLs showing epistatic effects only (epQTLs), and three occurred between M_epQTLs and epQTLs. Significant GE interactions were found for six QTLs with additive effects and one AA interaction. Generally, the contributions to the phenotypic variation were higher due to QTL main effects than to epistatic effects. The detection of additive effects and AA effects of a QTL interfered with each other, indicating that the detection of QTLs with main effects, as well as the magnitude and directions of the additive effects, might vary depending on their interactions with other loci.     

Stimulation of Na,K-ATPase by low potassium requires reactive oxygen species

The signaling pathway that transduces the stimulatory effect of low K⁺ on the biosynthesis of Na,K-ATPase remains largely unknown. The present study was undertaken to examine whether reactive oxygen species (ROS) mediated the effect of low K⁺ in Madin-Darby canine kidney (MDCK) cells. Low K⁺ increased ROS activity in a time- and dose-dependent manner, and this effect was abrogated by catalase and N-acetylcysteine (NAC). To determine the role of ROS in low-K⁺-induced gene expression, the cells were first stably transfected with expression constructs in which the reporter gene chloramphenicol acetyl transferase (CAT) was under the control of the avian Na,K-ATPase -subunit 1.9 kb and 900-bp 5'-flanking regions that have a negative regulatory element. Low K⁺ increased the CAT expression in both constructs. Catalase or NAC inhibited the effect of low K⁺. To determine whether the increased CAT activity was mediated through releasing the repressive effect or a direct stimulation of the promoter, the cells were transfected with a CAT expression construct directed by a 96-bp promoter fragment that has no negative regulatory element. Low K⁺ also augmented the CAT activity expressed by this construct. More importantly, both catalase and NAC abolished the effect of low K⁺. Moreover, catalase and NAC also inhibited low-K⁺-induced increases in the Na,K-ATPase ₁- and ₁-subunit protein abundance and ouabain binding sites. The antioxidants had no significant effect on the basal levels of CAT activity, protein abundance, or ouabain binding sites. In conclusion, low K⁺ enhances the Na,K-ATPase gene expression by a direct stimulation of the promoter activity, and ROS mediate this stimulation and also low-K⁺-induced increases in the Na,K-ATPase protein contents and cell surface molecules. Madin-Darby canine kidney cells; N-acetylcysteine; catalase     

NKG2D discriminates diverse ligands through selectively mechano‐regulated ligand conformational changes

Stimulatory immune receptor NKG2D binds diverse ligands to elicit differential anti‐tumor and anti‐virus immune responses. Two conflicting degeneracy recognition models based on static crystal structures and in‐solution binding affinities have been considered for almost two decades. Whether and how NKG2D recognizes and discriminates diverse ligands still remain unclear. Using live‐cell‐based single‐molecule biomechanical assay, we characterized the in situ binding kinetics of NKG2D interacting with different ligands in the absence or presence of mechanical force. We found that mechanical force application selectively prolonged NKG2D interaction lifetimes with the ligands MICA and MICB, but not with ULBPs, and that force‐strengthened binding is much more pronounced for MICA than for other ligands. We also integrated steered molecular dynamics simulations and mutagenesis to reveal force‐induced rotational conformational changes of MICA, involving formation of additional hydrogen bonds on its binding interface with NKG2D, impeding MICA dissociation under force. We further provided a kinetic triggering model to reveal that force‐dependent affinity determines NKG2D ligand discrimination and its downstream NK cell activation. Together, our results demonstrate that NKG2D has a discrimination power to recognize different ligands, which depends on selective mechanical force‐induced ligand conformational changes.     

The effect of 7,8,4´‐trihydroxyflavone on tyrosinase activity and conformation: Spectroscopy and docking studies

Tyrosinase is a ubiquitous enzyme that plays an essential role in the production of melanin. Effective inhibitors of tyrosinase have extensive applications in the medical, cosmetic and food industries. In this study, a combination of enzyme kinetics, ultraviolet (UV)‐visible absorption, fluorescence spectroscopic techniques and a computational simulation method was used to characterize the inhibitory mechanism of 7,8,4´‐trihydroxyflavone on tyrosinase. 7,8,4´‐Trihydroxyflavone was found to strongly inhibit the oxidation of l ‐DOPA by tyrosinase with an IC₅₀ value of 10.31 ± 0.41 μM. The inhibitory mechanism was determined to be reversible and non‐competitive with a K_i of 9.50 ± 0.40 μM. The UV absorption spectra showed that 7,8,4´‐trihydroxyflavone could chelate with copper ions and form a complex with tyrosinase. The intrinsic fluorescence of tyrosinase was quenched by 7,8,4´‐trihydroxyflavone through a static quenching mechanism. 7,8,4´‐Trihydroxyflavone was found to occupy a single binding site with a binding constant of 7.50 ± 1.20 × 10⁴ M^?1 at 298 K. The conformation of tyrosinase changed, and the microenvironment became more hydrophilic after 7,8,4´‐trihydroxyflavone binding. Thermodynamics parameters indicated that the binding was a spontaneous process and involved hydrogen bonds and van der Waals forces. The binding distance was evaluated to be 4.54 ± 0.05 nm. Docking simulation analysis further authenticated that 7,8,4´‐trihydroxyflavone could form hydrogen bonds with the residues His244 and Met280 within the tyrosinase active site. Our results will contribute to further understanding of the inhibitory mechanisms of 7,8,4´‐trihydroxyflavone against tyrosinase and will facilitate future screening for tyrosinase inhibitors.     

三江平原湿地——虎林地区夏季浮游植物群落结构特征

对三江平原湿地虎林地区水域的浮游植物群落结构进行了初步研究。在采集区域设置10个采样点,经鉴定共有133个浮游植物分类单位,隶属于8门10纲16目27科48属。该地区浮游植物群落组成以硅藻门(Bacillariophyta)、绿藻门(Chlorophyta)为主。各采样点浮游植物种类组成及细胞密度差异显著,采样点Ⅸ的浮游植物种类最丰富,采样点Ⅱ的浮游植物细胞密度最大。在三江平原湿地虎林地区发现了大量的β-中污指示种,经聚类和多维尺度分析评价,初步推断三江平原湿地虎林地区水域受到一定污染,呈中营养状态。