微生物遗传学与育种课程教学改革与探索

微生物遗传学与育种是“生物工程专业卓越人才实验班”和“生物工程国际留学生班”的必修课程。然而,传统授课模式在内容选择、教学方法及手段和考核形式等方面均存在诸多不足。为了提高教学质量和效果,促进天津科技大学微生物学教学领域的进步和发展,更加高效地培养国家需要、满足国际需求的创新领军人才,文中对微生物遗传学与育种的教学内容、教学方法及手段、课程考核方式进行了改革与探索。借助最新科研进展、课前预习体系、视频展示、考核方式多样化等形式对授课模式进行创新性改革。不仅使学生掌握了微生物遗传学与育种的相关专业知识,更加锻炼了学生的主观能动性、团队合作意识和专业外语表达水平,培养了学生对微生物遗传学相关科学知识的兴趣。     

Elovl5转基因果蝇产生更长链的脂肪酸

大多数昆虫体内的多不饱和脂肪酸 (Polyunsaturated fatty acids,PUFAs) 主要是碳链长为18个碳原子的多不饱和脂肪酸 (C18-PUFAs),几乎不含价值更高、生物活性更强的C20、C22等长链的多不饱和脂肪酸。文中利用黑腹果蝇Drosophila melanogaster (Fruit fly) 作为生物模型,将来源于小鼠的Δ6-脂肪酸延长酶Elovl5基因优化后转移到果蝇中使其表达。结果表明通过显微注射法成功将含有Elovl5基因的载体导入果蝇胚胎中,在荧光显微镜下检测到在果蝇整个发育阶段均有增强型绿色荧光蛋白 (Enhanced green fluorescent protein,EGFP) 表达,同时Elovl5基因的表达显著地促使果蝇体内C18多不饱和脂肪酸向着更长链多不饱和脂肪酸的生物合成方向转化。使用转基因技术得到体内富含C20、C22等长链多不饱和脂肪酸的转基因果蝇模型,将为进一步开展果蝇多不饱和脂肪酸生物合成的机制提供研究基础。     

南美天胡荽及其根际土壤水浸提液化感成分分析

为探讨南美天胡荽对其他植物种子萌发的影响以及筛选影响其他植物的主要化合物,该文采用种子萌发试验、气相色谱-质谱联用以及液相色谱-质谱联用的方法,分析了南美天胡荽不同溶剂浸提液对种子萌发的影响、南美天胡荽植株及其根际土壤浸提液成分。结果表明:(1)南美天胡荽不同溶剂浸提物均具有一定程度的抑制种子萌发作用。(2)气相色谱-质谱分析下,南美天胡荽植株水浸提液中共分离鉴定了35种化合物,其中,邻苯二甲酸二丁酯(15.2%)、10,15-十八烷二元酸(8.58%)、2,4-二叔丁基苯酚(6.81%)相对含量最高; 根际土壤水浸提液中共分离鉴定了17种化合物,其中,油酸酰胺(26.47%)、正二十七烷(9.63%)、十六酸乙酯(4.83%)相对含量最高。(3)液相色谱-质谱分析下,南美天胡荽植株水浸提液共分离鉴定了109种化合物,ESI⁺模式下,L-苯丙氨酸(3 483.99 ng·mg^-1)、木犀草素(2 306.64 ng·mg^-1)含量最多,ESI^-模式下,右旋奎宁酸(21 827.71 ng·mg^-1)、绿原酸(12 589.25 ng·mg^-1)含量最多; 根际土壤水浸提液中共分离鉴定了93种化合物,ESI⁺模式下,丁酸(7 660.53 ng·mg^-1)、棕榈酰胺(3 200.36 ng·mg^-1)含量最多,ESI^-模式下,正二十八酸(18 605.35 ng·mg^-1)、蔗糖(12 183.23 ng·mg^-1)含量最多。(4)南美天胡荽的潜在化感物质主要为脂肪酸类、酰胺类、酯类、芳香酸类化合物,而土壤中直接起化感作用的物质可能为丁酸、正二十八酸、羟基乙酸、油酸酰胺、棕榈酰胺、十六酸乙酯、苯甲酸,其中脂肪酸类化合物输入可能来源于南美天胡荽、土壤微生物和土壤动物,酰胺类、酯类、芳香类化合物则更可能来源于南美天胡荽植株。     

基于GC-MS和UPLC-QTOF/MS技术的灵芝孢子粉化学成分分析

采用极性不同的6种溶剂（石油醚、乙酸乙酯、丙酮、乙醇、甲醇和水）、按索氏提取法逐级萃取破壁灵芝孢子粉,并同时运用气相色谱-质谱联用（GC/MS）和超高效液相串联四极杆飞行时间质谱（UPLC-QTOF/MS）技术对各萃取物进行化学成分分析与鉴定。结果表明：GC/MS共鉴定出101种化合物,其中酸类10种、酯类40种、醇类7种、酮类6种、酚类2种、烃类18种、甾类9种和杂原子化合物9种;UPLC-Q-TOF/MS共推断出40种化合物,其中倍半萜类1种、二萜类1种、三萜类9种、生物碱类4种、酰胺类7种、有机酸类9种以及其他化合物9种。两种测定方法间共有化合物仅1种,仅存在于5种有机溶剂（石油醚、乙酸乙酯、丙酮、乙醇和甲醇）萃取物之一的化合物共105种,2种或2种以上萃取物共有的化合物共31种,实验方法较好地实现了样品中化合物组分的充分分离,扩大了可检测化合物的范围。研究结果为灵芝孢子粉中化学成分的系统分析与鉴定、及灵芝孢子粉的化合物谱图库的完善提供了基础资料,为相关药理、药效分析及灵芝的药用模式真菌研究提供参考。     

沙丁胺醇、布地奈德不同雾化吸入联合头孢哌酮钠舒巴坦钠对AECOPD患者血气指标和肺功能的影响

目的:探讨沙丁胺醇、布地奈德不同雾化吸入联合头孢哌酮钠舒巴坦钠对慢性阻塞性肺疾病急性加重期(AECOPD)患者血气指标和肺功能的影响。方法:选取本院2015年1月至2018年10月期间收治的210例AECOPD住院患者作为研究对象,随机分为对照组(头孢哌酮钠舒巴坦钠治疗)、持续雾化组(沙丁胺醇、布地奈德持续雾化治疗+头孢哌酮钠舒巴坦钠)、先后雾化组(先给予沙丁胺醇雾化、后给予布地奈德雾化治疗+头孢哌酮钠舒巴坦钠),每组各70例,对比三组的临床疗效、血气指标、肺功能及不良反应。结果:先后雾化组总有效率为91.43%,高于持续雾化组的78.57%及对照组的67.14%(P0.05)。先后雾化组患者治疗5天后氧分压(PaO_2)、氧饱和度(SaO_2)高于持续雾化组及对照组,且持续雾化组高于对照组(P0.05);先后雾化组患者治疗5天后二氧化碳分压(PaCO_2)低于持续雾化组及对照组,且持续雾化组低于对照组(P0.05)。先后雾化组患者第一秒用力呼气容积(FEV_1)、用力肺活量(FVC)、第一秒用力呼吸容积占用力肺活量的百分比(FEV_1/FVC%)、第一秒用力呼气容积占预计值百分比(FEV_1%)高于持续雾化组及对照组,且持续雾化组高于对照组(P0.05)。三组不良反应总发生率比较无统计学差异(P0.05)。结论:沙丁胺醇、布地奈德先后雾化吸入联合头孢哌酮钠舒巴坦钠治疗AECOPD患者疗效确切,可提高患者肺功能,改善血气相关指标。     

Legacy effects override soil properties for CO2 and N2O but not CH4 emissions following digestate application to soil

The application of organic materials to soil can recycle nutrients and increase organic matter in agricultural lands. Digestate can be used as a nutrient source for crop production but it has also been shown to stimulate greenhouse gas (GHG) emissions from amended soils. While edaphic factors, such as soil texture and pH, have been shown to be strong determinants of soil GHG fluxes, the impact of the legacy of previous management practices is less well understood. Here we aim to investigate the impact of such legacy effects and to contrast them against soil properties to identify the key determinants of soil GHG fluxes following digestate application. Soil from an already established field experiment was used to set up a pot experiment, to evaluate N₂O, CH₄ and CO₂ fluxes from cattle‐slurry‐digestate amended soils. The soil had been treated with farmyard manure, green manure or synthetic N‐fertilizer, 18 months before the pot experiment was set up. Following homogenization and a preincubation stage, digestate was added at a concentration of 250 kg total N/ha eq. Soil GHG fluxes were then sampled over a 64 day period. The digestate stimulated emissions of the three GHGs compared to controls. The legacy of previous soil management was found to be a key determinant of CO₂ and N₂O flux while edaphic variables did not have a significant effect across the range of variables included in this experiment. Conversely, edaphic variables, in particular texture, were the main determinant of CH₄ flux from soil following digestate application. Results demonstrate that edaphic factors and current soil management regime alone are not effective predictors of soil GHG flux response following digestate application. Knowledge of the site management in terms of organic amendments is required to make robust predictions of the likely soil GHG flux response following digestate application to soil.     

Footedness predicts escape performance in a passerine bird

Behavioral lateralization, which is associated with the functional lateralization of the two brain hemispheres, commonly exists in animals and can provide an individual with benefits such as enhanced cognition and dual tasking. Lateral bias in limb use, as a type of behavioral lateralization, occur in many species, but the reasons for the coexistence of left‐ and right‐biased individuals in a population remain poorly understood. We examined the footedness of male yellow‐bellied tits (Pardaliparus venustulus) when they used feet to clamp mealworms against a perch, and tested its association with other fitness‐related behavioral traits (i.e., feeding efficiency, exploration tendency, and escape performance). We expected differently footed individuals to have respective advantages in these behaviors and thereby coexist (“respective advantage” hypothesis). We found their footedness repeatable, and there was no population‐level bias. While no associations of feeding efficiency and exploration tendency with footedness were detected, the right‐footed individuals were found to be harder to catch than the other individuals. Future studies need to investigate the reasons for the right‐footed individuals' superior escape performance. Moreover, the escape advantage for being right‐footed and the lack of population‐level bias in footedness in male yellow‐bellied tits suggest that the benefits related to left footedness also remain to be explored.     

In vitro anti-oxidant activity, fluorescence quenching study and structural features of carbohydrate polymers from Phyllanthus emblica

The water-extracted carbohydrate polymers (WE) of Phyllanthus emblica are analyzed using chemical, chromatographic, and spectroscopic methods. Anion-exchange-chromatography of WE yielded four fractions (F1-F4) with different chemical compositions and all of them contain phenolics. The major fraction F4 possesses 50% polysaccharide and 26% phenol, and is a glycoconjugate. The antioxidant capacities of WE and F4 are comparable to standard anti-oxidants. Notably, activities of F1-F4 correlate with their phenol content. Evidence for the complexation of F4 with bovine serum albumin is presented by fluorescence quenching measurement. The results also indicate conformational change of protein at high carbohydrate polymer concentration.     

Use of 3-hydroxy fatty acid concentrations in a murine air pouch infection model as a surrogate marker for LPS activity: a feasibility study using environmental Burkholderia cenocepacia isolates

Using a murine hypodermic air pouch infection model designed to mimic the release of bacterial products at physiological levels, 3-hydroxy fatty acid (3-OH FA) and endotoxin unit levels from Burkholderia cenocepacia isolates were assessed. The B. cenocepacia environmental isolates (n = 35) survived in the hypodermic air pouch but did not invade across the peritoneal epithelial layer during a 72-h infection. For all 35 strains, when the molar ratio of C_14:0 3-OH FA to C_16:0 3-OH FA in the air pouch fluid wash samples was between 1.4 and 2.5, the concentrations of C_14:0 3-OH FA were correlated with the endotoxin unit levels. However, both surrogate markers exhibited different correlations to the inflammatory response. The linear regression coefficient was 0.4234 for C_14:0 3-OH FA concentrations vs. NO productions, 0.223 for endotoxin unit levels vs. NO productions, 0.5008 for C_14:0 3-OH FA concentrations vs. TNF-alpha productions and 0.2869 for endotoxin unit levels vs. TNF-alpha productions. Therefore, C_14:0 3-OH FA concentrations, rather than endotoxin unit levels, acted as an immunostimulatory indicator for LPS in the B. cenocepacia isolates.     

Effects of osmotic pretreatments on oxidative stress, antioxidant profiles and cryopreservation of olive somatic embryos

A three-day pretreatment of olive somatic embryos (SE) with 0.75 M sucrose, combined with cryoprotection (0.5 M DMSO, 1 M sucrose, 0.5 M glycerol and 0.009 M proline) and controlled rate cooling, supported regrowth (as 34.6% fresh weight gain) and resumption of embryo development after cryopreservation. Pretreatment with mannitol or sorbitol did not support regrowth. Profiles of sugars, proline, antioxidant enzymes, Reactive oxygen species (ROS), secondary oxidation products and ethylene were constructed for the most successful (0.75 M) pretreatment series. Sucrose was the optimal pretreatment for supporting recovery, it also elevated glutathione reductase (GR) activity compared to controls, whereas superoxide dismutase (SOD), catalase and guaiacol peroxidase activities remained relatively unchanged. Superoxide dismutase activity was higher in SE pretreated with sucrose, compared with those pretreated with polyols; H₂O₂ was enhanced in SE pretreated with sorbitol and sucrose compared to mannitol. The overall trend for ethylene and OH production revealed their levels were highest in SE pretreated with polyols albeit, for individual treatments this was not always the case. Generally, pretreatments did not significantly change embryo secondary oxidation profiles of ThioBarbituric Acid Reactive Substances (TBARS) and Schiff's bases. In combination these studies suggest oxidative processes may influence regrowth of cryopreserved olive SE and that optimal pretreatments could, in part, increase tolerance by an overall enhancement of endogenous antioxidants (particularly GR), proline and sugars.