Effects of calpain-2 and autophagy-related protein 5 on hepatocyte apoptosis induced by endoplasmic reticulum stress

AIMTo investigate the effects of calpain-2 and autophagy-related protein 5 (Atg5) on apoptosis of BRL-3A rat normal liver cells during endoplasmic reticulum stress (ERS) induced by dithiothreitol (DTT). METH?ODS: BRL-3A cells were treated with DTT at 2.0 mmol/L for 0, 6, 12 and 24 h to induce ERS. Real-time cell analysis (RTCA) was used to measure the effect of DTT on BRL-3A cell proliferation. Apoptosis and cell cycle distribution were analyzed by flow cytometry. The mRNA expression of calpain-2 and Atg5 was detected by real-time PCR. The protein levels of calpain-2, Atg5, Atg7, Atg12 and microtubule-associated protein 1 light chain 3 (LC3) were determined by Western blot. The interaction between calpain-2 and Atg5 was investigated by co-immunoprecipitation (Co-IP). RESULTSThe proliferation of BRL-3A cells treated with DTT was significantly inhibited. The apoptosis of BRL-3A cells was significantly increased after DTT treatment for 6, 12 and 24 h as compared with 0 h group (P<0.05). The cell cycle was arrested in G₁ phase after DTT treatment (P<0.05). After DTT treatment for 6, 12 and 24 h, the mRNA expression of calpain-2 and Atg5 in the BRL-3A cells was significantly increased as compared with 0 h group (P<0.05). The protein levels of calpain-2, Atg12 and Atg7 in the cells treated with DTT for 6, 12 and 24 h were significantly higher than those in 0 h group, and the ratio of LC3-II/LC3-I was also significantly higher than that in 0 h group, while Atg5 expression was significantly lower than that in 0 h group (P<0.05). The results of Co-IP found that the anti-calpain-2 antibody precipitated Atg5 protein from the cell lysates, and the anti-Atg5 antibody also precipitated calpain-2 from the cell lysates, which confirmed the interaction between calpain-2 and Atg5. CONCLUSION Calpain-2 may participate in ERS-induced hepatocyte apoptosis by interacting with Atg5.     

大花君子兰叶绿体基因组及其特征

采用Illumina MiSeq测序平台对大花君子兰（Clivia miniata）叶片总DNA进行测序，通过组装获得了其叶绿体基因组（cpDNA）全长序列（158 114 bp）。对其cpDNA注释得到135个基因，包含87个蛋白编码基因、40个tRNA基因和8个rRNA基因。采用生物信息学方法对获得的cpDNA进行简单序列重复（SSR）分析和密码子偏好性分析。结果显示：①大花君子兰cpDNA中共有61个SSR位点，其中单核苷酸、二核苷酸、三核苷酸、四核苷酸、五核苷酸和六核苷酸重复数分别为38、9、2、8、3和1个，多数SSR分布在基因间隔区；②大花君子兰cpDNA密码子偏爱以A或U（T）结尾，亮氨酸使用频率最高，半胱氨酸使用频率最低。基于24种植物的cpDNA全长和23种植物的叶绿体ycf2基因序列进行系统发育分析，结果显示大花君子兰与石蒜科植物在同一分支，显示最近的亲缘关系，支持大花君子兰属于石蒜科。基于叶绿体ycf2的系统发育分析结果与基于cpDNA全长的系统发育分析研究结果大部分相同，支持ycf2基因可以代替cpDNA全长用于植物系统发育分析。     

高花青素洋葱品种"秀球赤玉"的选育与特征特性

“秀球赤玉”是以高花青素深紫皮不育系562A为母本，以高花青素深紫皮自交系2014-7-10-6为父本培育的中日照、早熟、高花青素三系杂交种。其表现：鳞茎大圆球形，花青素含量高达68.8 mg/g；洋葱鳞片从外至里均为深紫色，色泽亮，外观漂亮，商品率高；辣味轻，口感脆甜，可生食；植株长势强，抗病，耐抽薹，分球、裂球少；假茎细且收口紧，耐贮性好；平均单球质量600～1 000 g，667 m2产量8 000～12 000 kg；适宜在中日照地区推广，2019—2020年累计推广面积超过1 500 hm2。     

不同光质下外源锌对水培生菜生长和品质的影响

以'意大利耐抽薹'生菜(Lactuca sativa L.)为试材,采用水培法,设置不同光质(红蓝光比例为4∶1、1∶2和全红光、全蓝光)和外源锌浓度(0、0.5、1.0 mg·L-1 ZnSO4·7H2O),研究不同光质下外源锌对水培生菜生长和品质特性的调控效应,以期为植物工厂在不同光质下施加外源锌提高蔬菜产量和品质提供参考依据.结果 表明:高比例红光会促进生菜生物量、可溶性糖和还原糖的积累,并降低硝酸盐的含量,其中以红蓝光比例为4∶1的光质处理效果最好;高比例蓝光会促进光合色素和可溶性蛋白质含量的增加,其中以全蓝光处理效果最好,但全蓝光处理会对植株的生长产生抑制作用.从光质与Zn的互作效果看,光质对生菜生长和品质的影响起主导作用,外源Zn起调节作用.综合考虑,红蓝光比例为4∶1光质下根施0.5 mg·L-1 ZnSO4·7H2O处理对生菜的生长发育、产量提高以及品质改善的促进作用最佳.     

屋顶绿化5种常绿植物的抗旱性研究

为了筛选更多耐粗放管理且抗旱性强的屋顶绿化常绿植物，选取在西安露地广泛应用且须根发达无主根的5种常绿灌木:金叶女贞、金边黄杨、南天竹、海桐、红叶石楠进行干旱胁迫，通过观察植株外部形态特征的变化，并对其叶片的生理指标:相对含水量、细胞质膜透性、叶绿素、丙二醛、脯氨酸的含量进行测定，利用隶属函数值法进行综合评价。结果表明，5种常绿灌木的抗旱性排序依次为金边黄杨＞海桐＞红叶石楠＞金叶女贞＞南天竹，应用于屋顶绿化时，金叶女贞、南天竹应加强灌溉措施。总体而言说明5种常绿灌木基本能适应西安市的气候条件和屋顶环境。     

Effects of different long-term cropping systems on phosphorus adsorption and desorption characteristics in red soils

Journal of Soils and Sediments - Phosphorus (P) fertilizer has been applied in regions with red soils to ensure high crop yield. However, the supply of bioavailable P for crop plants is...     

野猪粪便中乳酸菌的分离鉴定及特性研究

【目的】筛选安全的、具有优良特性的乳酸菌菌株,进一步研发益生制剂,为饲料添加剂等动物相关产品提供资源。【方法】从我国黑龙江省大兴安岭地区采集野猪粪便样品13份,编号后置于4℃保温箱迅速运回实验室,利用MRS培养基分离纯化乳酸菌。使用细菌基因组DNA提取试剂盒提取分离菌株的基因组DNA,利用细菌16S rDNA通用引物进行PCR鉴定,将扩增得到的序列测序后在NCBI上使用BLAST与GenBank数据库中序列进行对比分析,确定各菌株的分类学地位。将鉴定后的乳酸菌菌株分别接种于酸性（pH 3.0）和含胆盐（0.3%）的MRS培养基,在不同条件下评价乳酸菌菌株的耐酸、耐胆盐特性。将过夜培养的乳酸菌于室温条件下静置,在不同时间测定其OD_600nm,进行自凝集能力评价;过夜培养的菌株分别与致病性埃希氏大肠杆菌、金黄色葡萄球菌和鼠伤寒沙门氏菌3种致病菌混合后于室温静置,进行共凝集能力检测。在体外,分别进行乳酸菌菌株对Caco-2细胞和IPEC-J2细胞的黏附能力测定,评价不同菌株的黏附能力。通过测定乳酸菌菌株对致病性埃希氏大肠杆菌、金黄色葡萄球菌和鼠伤寒沙门氏菌3种致病菌的抑菌环直径,评价分离菌株的抑菌活性。通过体内外试验评价乳酸菌菌株的安全性。在体外,分别以模式菌株嗜酸乳杆菌和金黄色葡萄球菌作为阴性对照和阳性对照,将3株乳酸菌菌株和对照菌株在血平板上划线,37℃厌氧孵育18—24 h,观察细菌菌落周围是否形成溶菌环,评价分离菌株的溶血特性。使用文献中已报道的毒力因子引物对分离的乳酸菌菌株进行PCR扩增,检测是否存在毒力因子的编码基因,评估分离菌株的安全性。在体内,将过夜培养的乳酸菌连续饲喂7周龄的BALB/c小鼠21 d,分别测定小鼠的初始体重和最终体重,观察计算体重变化情况;饲喂21 d后,解剖获取小鼠的脾脏、肝脏和肾脏计算器官指数,评价分离乳酸菌菌株的体内安全性。【结果】从野猪粪便中分离得到3株对酸和胆盐具备一定耐受力的乳酸菌,经鉴定分别为蒙氏肠球菌（Enterococcus mundtii）、耐久肠球菌（Enterococcus durans）和黏膜乳杆菌（Lactobacillus mucosae）。3株乳酸菌菌株均表现出较强的自凝集能力和对致病菌的共凝集能力,同时对Caco-2细胞和IPEC-J2细胞均表现出较强的黏附能力,抑菌试验结果显示黏膜乳杆菌对3种致病菌均具备较强的抑菌活性。经体内外安全性评价,3株乳酸菌菌株无溶血性,且均未检测到毒力基因,经其连续饲喂的小鼠行为表现正常、状态良好,其中,与对照组相比,黏膜乳杆菌饲喂后小鼠增重显著。【结论】从大兴安岭野猪粪便中分离的3株乳酸菌（特别是黏膜乳杆菌）具有良好的特性和安全性,具备进一步开发益生菌制剂的潜力。     

Effect of Bacillus cereus as a water or feed additive on the gut microbiota and immunological parameters of Nile tilapia

We evaluated the effects of Bacillus cereus, as an additive in water and feed, on the gut microbiota and immunological parameters of Nile tilapia (Oreochromis niloticus) fingerlings. Experiments were performed in tanks and net cages respectively. Experiment 1: Tilapia were housed in tanks for 42 days, and B. cereus was added to the water at 1.0 × 10⁴ cfu mL^?1 (Treatment 1) and 1.0 ×10⁵ cfu mL^?1 (Treatment 2) weekly. For the control, no probiotic was added. Experiment 2: Tilapia were housed in cages for 42 days, and the feed was supplemented with B. cereus at 1.0 × 10⁷ cfu g^?1 (Treatment 1) and 1.0 × 10⁸ cfu g^?1 (Treatment 2) weekly. For the control, no probiotic was added. Each treatment contained three replicates, with 50 male tilapias per replicate. The fish from the probiotic treatments in both tank and cage experiments had significantly higher serum lysozyme and peroxidase activities than the control. In the cage experiment, alkaline phosphatase and total superoxide dismutase activities in tilapia were significantly higher in probiotic treatments compared with the control. The results of polymerase chain reaction‐denaturing gradient gel electrophoresis showed that B. cereus supplementation in the feed and water affected the autochthonous gut bacteria community of tilapia and stimulated various potentially beneficial bacteria. Therefore, B. cereus, as a water or feed additive, could enhance the immune status and affect the gut microbiota of tilapia. Bacillus cereus was more effective as a feed supplement rather than a water additive for enhancing the immune status of tilapia.     

Use of Microsatellite DNA Profiling to Identify Japanese Flounder,Paralichthys olivaceus of Hatchery Origin

Japanese flounder, Paralichthys olivaceus, is an economically important marine fish species in Asia. A suite of 18 microsatellite markers chosen from published genetic linkage maps was used to carry out parentage assignments of 188 hatchery‐reared juveniles from a small number of breeders. The probabilities of exclusion for the 18 microsatellite markers were 0.604–0.913, and the effectiveness of combined probability of exclusion reached 100% when using the eight microsatellite markers with higher Excl 1 probabilities. The cultured and wild stocks (WSs) were differentiated in a release‐recapture population based on these markers. Of the 321 recaptured offspring, 28.34% were assigned to their parental pairs in our broodstock, whereas the remaining offspring could not be traced back to a possible sire or dam. Significant reduction in genetic diversity of the cultured stock (CS) had not been found compared with that of the WS. The results suggest that CSs released into the wild will not adversely affect the genetic structure of natural populations. Our results demonstrate that these markers provide an efficient tool for parentage assignments and genetic analysis of Japanese flounder.     

Dietary fishmeal levels affect the volatile compounds in cooked muscle of farmed large yellow croaker Larimichthys crocea

A comparative study on the volatile compounds in cooked muscle of wild and farmed large yellow croaker (LYC) was conducted. The two farmed LYC groups were fed with diets containing 44% (CF) and 25% (LF) of fish meal (FM) respectively. Results showed that 48 volatiles, including aldehydes, alcohols, ketones, hydrocarbons, aromatics, acids, esters, furans and miscellaneous compound, were detected in cooked fillets. The LF group had significantly lower amounts of total aldehydes and ketones, higher content of miscellaneous compound in cooked fillets than that in the CF and wild groups (p < .05). Compared with the wild group, the LF group had significantly lower amounts of total alcohols, acids and esters, while the CF group had significantly lower amounts of total aldehydes, higher content of total ketones in cooked muscle (p < .05). According to the principal component analysis (PCA), some volatiles (propanal, nonanal, etc.) could be considered as sensitive indicators to classify cooked muscle samples. In conclusion, differences in the volatiles in the cooked muscle between the wild and farmed LYC have been found. Low level of dietary FM (25%) changed the volatile profiles in cooked fillets of farmed LYC. A PCA may be useful to screen potential volatiles to classify cooked muscle samples in this study.