Using vibrational molecular spectroscopy to detect moist heating induced carbohydrates structure changes in cool-climate adapted barley grain

There is limited research to study how moist heating affects internal structure of barley grain on a molecular basis. The objectives of this study were to use vibrational molecular spectroscopy: 1) to determine the moist heating induced changes of barley carbohydrate (CHO) structure on a molecular basis, 2) to study the effects of moist heating on CHO chemical profiles, Cornell Net Carbohydrate and Protein System (CNCPS) subfractions, in situ rumen degradation, and predicted intestinal carbohydrate supply of barley grain; and 3) to reveal the association between molecular structure spectral features and CHO related metabolic characteristics. Barley samples (CDC cowboy) were collected from Kernen Crop Research Farm (Saskatoon, Canada) during two consecutive years. Half of each sample was kept as raw barley and the other half underwent moist heating (autoclaving at 120 °C for 60 min). The molecular spectroscopy (attenuated total reflectance-fourier transform infrared, ATR-FTIR) was used to detect the barley CHO related molecular structure spectral features. Moist heating did not affect carbohydrate related chemical profiles and CNCPS subfractions but it decreased rumen degradable carbohydrate. Rumen undegradable and intestinal digestion of CHO subfractions were not affected by moist heating. The advanced vibrational molecular spectroscopy can be used to detect carbohydrate molecular spectral features. Nutrient utilization prediction using molecular spectral characteristics is warranted and further investigation is encouraged.     

Growing Degree Day: Noninvasive Remotely Sensed Method to Monitor Diet Crude Protein in Free-Ranging Cattle

The daily nutritional balance of free-ranging cattle is the net result of intake from available forage biomass and nutritive value weighed against the nutritional requirements of the animal. Plant phenology influences nutritive value. Plant phenology is dictated by time of year and an accumulation of photosynthetically active days. Growing degree day (GDD) is a concept that quantifies this relationship and has been used to predict nutritive value in perennial range grasses. GDD could be substituted for chemical analysis to inform grazing animal nutritional monitoring efforts. We hypothesized that in C4 grass-dominated rangelands, a cumulative GDD calculation would correlate with diet crude protein (CP) predictions obtained by fecal near infrared spectroscopy (FNIRS) from free-ranging cattle. Therefore, the objectives of our research were to evaluate the effectiveness of GDD to predict FNIRS-derived determinations of grazing cattle diet CP in 1) two groups of three individual animals grazing a small native pasture and 2) large commercial-scale herds grazing expansive rangelands. For the first objective, cumulative GDD and FNIRS-predicted diet CP were strongly correlated (r² = 0.76; P < 0.01). Relationships between cumulative GDD and FNIRS-predicted diet CP for the second objective varied considerably among ranches, ranging from a low r² of 0.05 (P = 0.871) to a high r² of 0.78 (P < 0.049). Similar values for individual ranch/year combinations were stronger; ranging from a minimum r² of 0.44 (P = 0.556) to a maximum of 0.95 (P = 0.051). The aggregate relationship between GDD and FNIRS-predicted CP for all ranch/year combinations was highly significant (r² = 0.37; P < 0.001), but the standard error was 1.86% CP. The noninvasive remotely sensed grazing animal nutritional monitoring method described here was accurate enough to inform tactical rangeland diet quality assessments but was not accurate enough to inform operational-scale grazing management decisions.     

玉米瘤黑粉菌SG200效应蛋白PEP1表达及生物信息学分析

运用生物信息学手段分析pep1基因结构,并根据GenBank中玉米瘤黑粉菌pep1 DNA序列设计引物,从玉米瘤黑粉菌SG200的基因组中扩增得到了pep1基因全长,构建了pET–28a–pep1重组表达质粒,选用大肠埃希菌BL21(DE3)作为宿主菌,以1 mmol/L IPTG诱导表达。SDS–PAGE检测结果表明,诱导表达产物大小与理论值(20 800)一致,说明pET–28a–pep1能够在大肠埃希菌BL21(DE3)中高效表达。     

不同氮肥施用量对胶东卫矛氮代谢生理影响的研究

分析氮对胶东卫矛氮代谢生理的影响规律,明确氮肥施用后的相关酶活性变化机理,为其育苗中科学施用氮肥提供理论依据。试验在田间条件下,设置氮肥施用量分别为0 g/株(J1,CK)、9 g/株(J2)、18 g/株(J3)、27 g/株(J4)4个处理,3次重复。结果表明：5—9月J3谷草转氨酶活性分别比J1提高了64.84%、70.77%、58.52%、47.33%、57.20%,J4与J3之间无显著差异;6—9月J3谷丙转氨酶活性分别比J2提高了18.00%、30.79%、16.47%、16.83%,差异显著;谷胺酰胺合成酶活性7、9月J3显著高于J2,5—6月J2、J3、J4之间无显著差异,J2显著高于对照;5—9月J3硝酸还原酶活性分别比J2提高了20.47%、31.66%、27.32%、23.50%、34.90%,差异显著,J3与J4之间无显著差异;5—7月J3硝态氮含量分别比对照提高了73.76%、53.90%、85.95%,差异显著,8—9月J2、J3、J4之间无显著差异;6—8月,J3可溶性蛋白含量分别比J2提高了15.90%、20.09%、9.04%,差异显著。综合分析认为,J3处理可以显著提高胶东卫矛氮代谢相关酶活性,提高植株叶片内硝态氮含量和可溶性蛋白含量,效果优于J2、J4处理。     

小麦热激蛋白转录因子C家族基因的结构及其在干旱胁迫中的表达

小麦热激蛋白转录因子在热胁迫和耐热性产生的过程中发挥重要作用。为进一步了解小麦热激蛋白转录因子C亚家族,本研究采用全基因组信息保守结构域比对和进化聚类分析,共鉴定了24个小麦TaHsfC基因,并对TaHsfC亚家族成员染色体定位、基因结构、亚细胞定位、对应蛋白质的氨基酸特点、基因表达进行了分析。结果表明,24个TaHsfCs主要分布在5条染色体上,其中A基因组中有7个,B基因组中9个,D基因组中有6个,另外两个所在染色体位置不清楚。TaHsfCs含有0～2个内含子,其中 TaHsfC1亚族基因含有1～2个内含子, TaHsfC2亚族基因则含有0～1个内含子。聚类分析表明,小麦TaHsfC成员共分为 TaHsfC1和 TaHsfC2两个亚族。24个TaHsfCs成员全部定位于细胞核。在15%PEG模拟干旱条件下,干旱敏感品种矮抗58和耐旱品种晋麦47中有10个TaHsfC基因显著上调表达,其中 TaHsfC2亚族基因的表达量上调倍数均高于 TaHsfC1亚族基因(其中 TaHsfC2j和 TaHsfC2k未在晋麦47中检测到表达量)。本研究可为探索小麦热激蛋白转录因子C家族基因在小麦抗旱中的分子机制提供一定的理论支撑。     

三种酶制备牡蛎蛋白水解物的抗氧化活性和功能特性分析

目的：比较分析木瓜蛋白酶、胃蛋白酶和胰蛋白酶三种不同酶水解牡蛎获得的蛋白产物抗氧化活性和功能特性。方法：采用木瓜蛋白酶、胃蛋白酶和胰蛋白酶三种不同酶水解牡蛎蛋白获得水解物,并分别测定它们的在不同pH下的溶解度、乳化性以及不同浓度水解物的还原能力、DPPH自由基清除率和Fe^{2 +}螯合能力。结果：表明三种酶水解物都有较高的溶解度(p＜0.01),胰蛋白酶产生的水解物溶解高度达94.8％(p＜0.05)；水解物溶解度和乳化活性指数在pH4显著降低(p＜0.05)。牡蛎蛋白质水解产物的还原能力、DPPH自由基清除能力和金属螯合活性随浓度的增加而增强,其中胰蛋白酶和胃蛋白酶抗氧化能力较强(p＜0.01)。结论：三种蛋白酶水解牡蛎蛋白可获得抗氧化活性强、溶解度高的水解物,并受浓度和pH的影响,它们的水解物可作为一种潜在的抗氧化功能多肽利用。     

草鱼TAB1蛋白的原核重组表达及其抗体制备

为了制备草鱼重组TAB1蛋白(rCiTAB1)及其特异性抗体,首先以草鱼头肾组织c DNA为模板,PCR扩增Ci TAB1基因全长序列,并依次构建重组克隆质粒p MD19-T-Ci TAB1与重组表达质粒pET-32a-Ci TAB1。该重组表达质粒经0.5 mmol·L~(-1) IPTG,37℃诱导表达12 h后获得以包涵体形式表达的重组CiTAB1蛋白(rCiTAB1)。然后,采用3种不同方法对包涵体蛋白进行变性,发现与高浓度尿素直接变性法和洗涤后尿素变性法相比,洗涤后梯度尿素变性法处理后的蛋白纯度最好,经透析复性后得到浓度为2 mg·mL~(-1)的r Ci TAB1。最后,将rCiTAB1蛋白与白油佐剂及免疫增强剂混合,室温下混合1.5h乳化成免疫原,3次免疫新西兰大白兔制备兔抗CiTAB1抗体,经间接ELISA方法和免疫琼脂双向扩散试验测得免疫后第33天血清中特异性抗体的效价分别为1∶1 048 576和1∶16。Western blot检测到一条分子量约为72 kDa的特异性条带,表明该抗体能特异性识别r Ci TAB1蛋白。研究结果为后续深入研究CiTAB1蛋白功能提供了物质基础。     

SIRT1 promotes autophagy of pancreatic cancer cells induced by hypoxia via regulating FOXO1/RAB7 signaling pathway

AIM: To investigate the effect of SIRT1 on the autophagy of pancreatic cancer cells under hypoxia condition, and to analyze the underlying mechanism of regulating FOXO1/RAB7 signaling pathway. METHODS: Western blot and immunofluorescence methods were used to determine the expression of SIRT1 in the pancreatic cancer cells. The small interfering RNA targeting SIRT1 and SIRT1 over-expression plasmid were transfected into the pancreatic cancer Panc-1 cells. Confocal microscopy was used to detect the LC3 expression. Western blot was used to analyze the protein levels of LC3, p62 and FOXO1/RAB7 signaling pathway-related molecules. Co-immunoprecipitation was used to detected the protein interaction between SIRT1 and FOXO1. RESULTS: The expression level of SIRT1 in the nucleus of Panc-1 cells was increased under hypoxia condition. Compared with negative control under hypoxia condition, knock-down of SIRT1 expression attenuated the autophagy flux in the pancreatic cancer Panc-1 cells (P<0.05). Over-expression of SIRT1 increased the protein levels of FOXO1 and RAB7. On the contrary, knock-down of SIRT1 expression inhibited the protein levels of FOXO1 and RAB7. The protein interaction between SIRT1 and FOXO1 in the pancreatic cancer cells was observed. CONCLUSION: SIRT1 in pancreatic cancer Panc-1 cells under hypoxia condition is over-expressed in the nucleus. Down-regulation of SIRT1 inhibits autophagy and its mechanism may be related to FOXO1/RAB7 signaling pathway.     

Expression and mechanisms of hsa_circ_087631 in patients with primary biliary cholangitis

AIM To explore the expression and mechanisms of circular RNA hsa_circ_087631 in the patients with primary biliary cholangitis （PBC）. METHODS RT-qPCR was used to detect the expression of hsa_circ_087631 in PBC patients and healthy controls. Hsa-miR-346-overexpressing lentiviral vector pLenti-EF1a-EGFP-F2A-Puro-CMV-MCS was constructed and transfected into human acute T cell leukemia Jurkat cells， and then the expression levels of hsa_circ_087631， Bcl-6 mRNA and interleukin-21 （IL-21） mRNA were detected by RT-qPCR. Dual-luciferase reporter assay was performed to identify the interactions between hsa_circ_087631 and hsa-miR-346. RESULTS The expression of hsa_circ_087631 in the PBC patients was significantly increased compared with the healthy subjects. Hsa-miR-346-overexpressing lentiviral vector pLenti-EF1a-EGFP-F2A-Puro-CMV-MCS was successfully constructed. The expression of hsa-miR-346 was significantly increased after the hsa-miR-346-overexpressing lentiviral vector was transfected into the Jurkat cells， while the expression levels of hsa_circ_0087631， Bcl-6 mRNA and IL-21 mRNA were significantly decreased. After wild-type or mutant hsa_circ_087631 vector and hsa-miR-346 mimics were transfected into 293T cells， the results of dual-luciferase reporter assay showed that hsa-miR-346 significantly decreased the luciferase activity of wild-type hsa_circ_087631 （P<0.01）， but the regulation did not change significantly after mutation of the predicted binding site. CONCLUSION Peripheral blood hsa_circ_087631 level is elevated in the PBC patients. The hsa_circ_087631/hsa-miR-346/Bcl-6 signaling may take effect in human T cells. Hsa-miR-346 significantly reduces the expression of hsa_circ_087631， but it may not be regulated by predicted binding sites.