两种牙体充填材料对乳牙边缘微渗漏及固化效率的体外研究

目的:比较纳米瓷充填材料和复合树脂充填乳牙V类洞边缘微渗漏的影响以及固化效率的差异。方法:第一部分为微渗漏实验,选取滞留拔除的上颌乳前牙20颗,唇侧制备V类洞,随机分为A、B两组,分别用Composan LCM和Filtek~(TM) Z350充填,亚甲基蓝染色处理,24 h后使用体式显微镜观察,测量染料渗入深度为微渗漏值,每组各选2个样本扫描电镜观察充填体与牙体组织间粘结情况。第二部分为显微硬度实验,分别用两种充填材料制作模块各9个,记为A、B两组,材料模块光固化24 h后使用显微硬度仪检测充填材料的固化效率。结果:Composan LCM组微渗漏值明显低于Filtek~(TM)Z350组(P0.05);扫描电镜下观察Filtek~(TM) Z350组充填材料与牙体组织间可见裂缝,界面参差不齐,有不连续的小断裂,Composan LCM组界面连续均匀,未见明显裂缝;两组充填材料的固化效率无显著差异(P0.05)。结论:纳米瓷充填材料和复合树脂均可用于乳牙龋洞充填,使用纳米瓷充填材料可减少微渗漏。     

宫颈特殊染色法、液基薄层细胞学及人乳头瘤病毒检测对宫颈癌前病变的应用价值分析

目的:探讨宫颈特殊染色法(FRD)、液基薄层细胞学(TCT)及人乳头瘤病毒(HPV)检测对宫颈癌前病变筛查的应用价值。方法:选取2015年1月～2018年1月于我院行宫颈癌筛查的1794例妇女作为研究对象,所有研究对象均接受FRD、TCT、HPV检测,以经阴道镜取样活检结果为阳性标准,对比分析三种不同检测方法以及联合检测的诊断效能。结果:病理科活检检出阳性111例,检出率为6.19%;FDR检测检出阳性114例,检出率为6.35%,漏诊率为16.22%;TCT检测检出阳性115例,检出率为6.41%,漏诊率为19.82%;HPV检测检出阳性108例,检出率为6.02%,漏诊率为19.82%;FRD检测与TCT、HPV检测的检出率比较差异无统计学意义(P0.05)。FRD检测敏感度为83.78%,特异度为98.75%,阳性预测值为81.58%,阴性预测值为98.93%;TCT检测敏感度为80.18%,特异度为98.46%,阳性预测值为77.39%,阴性预测值为98.69%;HPV检测敏感度为80.18%,特异度为98.87%,阳性预测值为82.41%,阴性预测值为98.70%;FRD、TCT、HPV联合检测敏感度为93.69%,特异度为99.52%,阳性预测值为92.86%,阴性预测值为99.58%;FRD、TCT、HPV联合检测与FRD、TCT、HPV单独检测的敏感度、阳性预测值比较差异具有统计学意义(P0.05)。结论:FRD、TCT、HPV检测对宫颈癌前病变的诊断效能相当,而FRD、TCT、HPV联合检测的诊断效能优于各方法单独检测。     

玻璃化冻存延期回植技术在大面积皮肤撕脱伤中的应用研究

目的:探讨玻璃化冻存延期回植技术用于大面积撕脱皮肤伤的临床效果。方法:选择2015年6月至2018年6月收治的大面积皮肤撕脱伤的患者16例,根据处理方式分为撕脱皮肤玻璃化冻存二期回植组和撕脱皮肤一期回植组,每组8例患者。皮肤冻存后回植前,采用MTT法检测皮肤活力,比较两组皮片回植后的成活率、感染率、平均住院时间。结果:玻璃化冻存组皮肤冻存后平均活力为92.56%±7.20%,皮片成活率为81.74%±5.78%,感染率12.5%,平均住院时间33.13±3.91 d;一期皮肤回植组皮片成活率21.91%±5.28%,感染率100%,平均住院时间47.50±1.93 d。玻璃化皮肤保存组皮片成活率、感染率、平均住院时间均优于一期皮肤回植组(P0.05)。结论:皮肤玻璃化冻存延期回植技术用于无法一期回植皮肤撕脱伤的临床效果较好。     

食品级乳酸乳球菌pNZ8149-luc质粒的构建及表达

目的:构建能够稳定表达萤火虫荧光素酶报告基因(luc)的乳酸乳球菌(Lactococcus lactis, L.lactis)食品级表达系统,以便后续研究对目的基因进行示踪。方法:从pGL4.10质粒中PCR扩增萤火虫荧光素酶报告基因,测序,克隆至载体pNZ8149,构建pNZ8149-luc表达质粒;电击转化宿主乳酸乳球菌NZ3900,采用乳糖筛选法获得重组的乳酸乳球菌,Nisin诱导,采用微孔板发光检测仪检测荧光素酶的存在,Western Blot检测目标蛋白luc的表达。结果:PCR扩增的荧光素酶报告基因成功克隆至pNZ8149质粒,并电击转化宿主乳酸乳球菌NZ3900,得到乳酸乳球菌表达系统NZ3900/pNZ8149-luc。Nisin诱导后,检测到荧光素酶随诱导时间的延长活性逐渐增强,时间超过24 h之后荧光素酶活性逐渐下降。Western Blot检测到目标蛋白luc在胞内表达。结论:成功构建了p NZ8149-luc表达载体,并能够在乳酸乳球菌体内稳定表达。     

Structural and Functional Study of d-Glucuronyl C5-epimerase

Heparan sulfate (HS) is a glycosaminoglycan present on the cell surface and in the extracellular matrix, which interacts with diverse signal molecules and is essential for many physiological processes including embryonic development, cell growth, inflammation, and blood coagulation. d-Glucuronyl C5-epimerase (Glce) is a crucial enzyme in HS synthesis, converting d-glucuronic acid to l-iduronic acid to increase HS flexibility. This modification of HS is important for protein ligand recognition. We have determined the crystal structures of Glce in apo-form (unliganded) and in complex with heparin hexasaccharide (product of Glce following O-sulfation), both in a stable dimer conformation. A Glce dimer contains two catalytic sites, each at a positively charged cleft in C-terminal α-helical domains binding one negatively charged hexasaccharide. Based on the structural and mutagenesis studies, three tyrosine residues, Tyr⁴⁶⁸, Tyr⁵²⁸, and Tyr⁵⁴⁶, in the active site were found to be crucial for the enzymatic activity. The complex structure also reveals the mechanism of product inhibition (i.e. 2-O- and 6-O-sulfation of HS keeps the C5 carbon of l-iduronic acid away from the active-site tyrosine residues). Our structural and functional data advance understanding of the key modification in HS biosynthesis.     

Functional expression of a novel alkaline-adapted lipase of Bacillus amyloliquefaciens from stinky tofu brine and development of immobilized enzyme for biodiesel production

Using enrichment procedures, a lipolytic strain was isolated from a stinky tofu brine and was identified as Bacillus amyloliquefaciens (named B. amyloliquefaciens Nsic-8) by morphological, physiological, biochemical tests and 16S rDNA sequence analysis. Meanwhile, the key enzyme gene (named lip _BA) involved in ester metabolism was obtained from Nsic-8 with the assistance of homology analysis. The novel gene has an open reading frame of 645 bp, and encodes a 214-amino-acid lipase (Lip_BA). The deduced amino acid sequence shows the highest identity with the lipase from B. amyloliquefaciens IT-45 (NCBI database) and belongs to the family of triacylglycerol lipase (EC 3.1.1.3). The lipase gene was expressed in Escherichia coli BL21(DE3) using plasmid pET-28a. The enzyme activity and specific activity were 250 ± 16 U/ml and 1750 ± 153 U/mg, respectively. The optimum pH and temperature of the recombinant enzyme were 9.0 and 40 °C respectively. Lip_BA showed much higher stability under alkaline conditions and was stable at pH 7.0–11.0. The Km and Vmax values of purified Lip_BA using 4-nitrophenyl palmitate as the substrate were 1.04 ± 0.06 mM and 119.05 ± 7.16 μmol/(ml min), respectively. After purification, recombinant lipase was immobilized with the optimal conditions (immobilization time 3 h at 30 °C, with 92 % enzyme recovery) and the immobilized enzyme was applied in biodiesel production. This is the first report of the lipase activity and lipase gene obtained from B. amyloliquefaciens (including wild strain and recombinant strain) and the recombinant Lip_BA with the detailed enzymatic properties. Also the preliminary study of the transesterification shows the potential value in biodiesel production applications.     

高效薄层色谱测定湛江等鞭金藻培养过程脂质变化

【目的】研究湛江等鞭金藻(Isochrysis zhangjiangensis)由氮元素丰富至限制培养过程中脂质成分的变化。【方法】利用高效薄层色谱(HPTLC)分离分析微藻中脂质。【结果】随着培养基中营养物质的消耗,细胞逐渐处于胁迫状态,在这种状态下,细胞大量积累贮存脂质—甘油三脂(TAG),组成生物膜系统的单半乳糖甘油二脂(MGDG)、硫代异鼠李糖甘油二脂(SQDG)、磷脂酰甘油(PG)和磷脂酰胆碱(PC)含量降低,而游离脂肪酸(FFA)、甘油二脂(DAG)、双半乳糖甘油二脂(DGDG)、磷脂酰肌醇(PI)和磷脂酰乙醇胺(PE)则相对稳定。【结论】HPTLC可作为一种简便、可靠的微藻中脂质分离分析方法,为研究微藻油脂代谢途径以及甘油三脂(TAG)的调控积累提供有效手段。     

过表达LeNLP4转录因子提高番茄抗低温胁迫能力

NAC（NAM-ATAF1,2-CUC2）转录因子在植物胁迫响应中起重要作用。为了探讨三舭丹基因在番茄抗低温胁迫中的功能,分离了番茄LeNLP4转录因子基因,并获得转正义LeNLP4基因番茄植株。荧光定量PCR分析表明,LeNLP4的表达受低温诱导。与野生型植株相比,在4℃胁迫下转基因植株具有较高的生长量和光系统II（PSH）最大光化学效率（Fv／Fm）、过氧化氢（H2O2）和超氧阴离子（O2-）清除速率、抗坏血酸过氧化物酶（APX）和超氧化物歧化酶（SOD）活性,以及较低的丙二醛（MDA）含量和相对电导率（REC）。过表达株系中SICBF1的表达高于野生型。上述结果表明,LeNLP4的过表达提高了转基因番茄抗低温胁迫能力。     

大豆食心虫储存蛋白hexamerin基因的克隆与表达分析

【目的】储存蛋白是昆虫发育、变态和生殖过程中氨基酸的主要来源,Hexamerin是储存蛋白家族重要成员,在昆虫的生长发育中起着重要作用。为了研究hexamerin基因(SpbHex)在大豆食心虫Leguminivora glycinivorella Matsumurav生长发育过程中的作用,对大豆食心虫SpbHex基因进行克隆与表达分析。【方法】利用RT-PCR和RACE技术克隆SpbHex的cDNA全长序列,并通过qPCR研究其在不同发育阶段和幼虫不同组织的表达情况。【结果】SpbHex基因cDNA序列全长2 161 bp,其中开放阅读框2 112 bp,编码703个氨基酸。蛋白预测分子量84.15 ku。hexamerin基因在大豆食心虫不同发育阶段和不同组织中均有表达。在不同生长发育阶段中4龄幼虫的表达量较高,1龄和成虫的表达量较低;在不同组织中脂肪体的表达量较高,表皮中的表达量最低。【结论】本研究克隆了大豆食心虫储存蛋白hexamerin基因,并对其在大豆食心虫中表达模式进行分析,为进一步明确hexamerin基因在大豆食心虫生长发育中的作用奠定基础。     

四霉素防治杨树溃疡病田间应用研究

杨树溃疡病是在杨树上发生普遍且危害严重的干部病害。为了有效控制杨树溃疡病的发生,利用四霉素制剂对杨树溃疡病进行田间防治试验。结果表明,于春季杨树溃疡病发生初期开始施药防治,采用刮涂法施药,0.3%四霉素水剂对杨树溃疡病有较好的防治效果,推荐使用量为有效成分60~300 mg/kg(稀释倍数10~50),防效为62.9%~77.4%。试验期间林间系统观察表明,0.3%四霉素水剂在试验浓度范围内对杨树生长发育无不良影响。