猪血红蛋白(Hb)脉冲酶解效果比较研究

目的寻找一种高效快捷有效地降解猪血红蛋白(Hb)新方法。方法在波型为双向方波,电极间距离为1.2 cm,脉冲频率为200 kHz的脉冲电场下,利用胰蛋白酶在温度为37℃,水解时间为4 h条件下水解猪血红蛋白。结果在脉冲电场作用下,胰蛋白酶水解血红蛋白获得的降解产物,利用高效凝胶色谱、紫外可见扫描及SDS-PAGE蛋白质电泳检测,发现其吸收峰或色带明显多于单一利用胰蛋白酶降解血红蛋白所得降解产物的吸收峰或色带。结论当脉冲电场通过血红蛋白时,血红蛋白内部的分子结构便产生斯塔克效应(Stark effect),引起血红蛋白分子剧烈振动,从而改变其分子结构振辐、吸收峰和偶极矩,并分别引起斯塔克频率、偶极矩、极化率的改变、使血红蛋白分子结构的极化跃迁和超极化,因此,在脉冲电场作用下,促进了血红蛋白酶解反应。     

DHPLC对猪肌肉组织差异表达EST的鉴定

采用DHPLC系统和Labwork4．0图像分析软件,对大白猪和广东地方品种蓝塘猪眼肌组织差异显示EST进行鉴定,两种方法均证实了差异显示条带的真实性。结果表明：用DHPLC系统能够准确而简捷的检测不同组织中mRNA表达的差异,是1种鉴定基因表达差异的有效方法,在研究基因表达,比较不同环境条件下动物组织的mRNA表达差异等方面具有广阔的应用前景。     

吸血诱导的埃及伊蚊海口株后期胰蛋白酶基因测序及与美国株同源性比较

应用逆转录-聚合酶链式反应（RT-PCR）技术从吸血后24 h埃及伊蚊海口株总RNA中扩增出了后期胰蛋白酶编码区cDNA序列。采用自动DNA分析仪进行序列分析,并与已知埃及伊蚊美国株后期胰蛋白酶基因及推导的氨基酸序列进行了同源性比较。结果表明：埃及伊蚊海口株后期胰蛋白酶基因序列与美国株同源性达98%,有11个碱基发生变异;氨基酸同源性达99%,仅有3个氨基酸发生变异,但与催化位点密切相关的氨基酸及N末端氨基酸序列完全一致。以上结果显示,埃及伊蚊胰蛋白酶不同地理株间存在微小的差异。     

MDA-DHPLC技术在PGS中的应用

目的:评估丙二醛变性高效液相色谱法在胚胎移植过程中的临床价值。方法:收集我院生殖中心2010年2月~2012年3间未做移植的废弃胚胎,由D2胚胎培养到D3胚胎期,提取遗传物质并进行PRC扩增,采用高形态学评分的D3作为对照,高效液相色谱法测定遗传DNA过氧化产物丙二醛(MDA)的含量。结果:多核Ⅰ组D2期卵裂球核≥2个,共24枚胚胎,DNA过氧化产物丙二醛(MDA)的含量5.32±0.19μmol/L,对比正常胚胎0.67±0.08μmol/l有显著差异(P0.05)。多核Ⅱ组D2期卵裂球核=2,共19枚胚胎,MDA的含量4.12±0.22μmol/L,对比正常胚胎0.67±0.08μmol/l有显著差异(P0.05)。Ⅰ组与Ⅱ组间没有统计学差异。然而,空泡胚胎与非空泡胚胎之间MDA并没有差异。结论:多核D2期胚胎培养到D3期会产生很高异常率,因此临床上应减少使用此种进行移植。     

一株产紫杉醇内生真菌液体发酵工艺的优化

紫杉醇是一种高效、低毒、广谱的天然抗癌药物,可以有效地治疗乳腺癌、子宫癌等。近年来的研究发现,从植物内生真菌中发酵生产紫杉醇被证明是解决药源问题的有效途径。从分离到的420株内生真菌中筛选到一株产紫杉醇的内生真菌XC1-07为实验材料进行发酵条件的初步优化。结果表明：最适碳源、氮源分别是麦芽糖和NH4NO3;在含10g/L NH4NO3、90g/L麦芽糖、1.0g/L MgSO4、pH6的优化培养基中培养13d,紫杉醇的产量为1,124.34μg/L。为目前报道的植物内生真菌发酵生产紫杉醇最高的产量。     

微生物燃料电池降解苯酚过程中微电场对阴极微生物多样性的影响

[背景]苯酚废水作为一种毒性强、难降解的废水而备受关注.目前,微生物燃料电池(microbial fuel cell,MFC)已经广泛用于苯酚废水的降解,MFC的产电效果和苯酚的降解效率与反应器内的微生物群落有着密切关系.[目的]为了提高MFC的产电效果及对有害物质的降解能力,需要对MFC中苯酚的降解和微生物群落结构进...     

基于UPLC-Q-Exactive技术结合OTCML数据库快速分析沙棘的功效成分

为探讨沙棘果茎叶功效成分的整体差异。实验应用超高效液相色谱-四极杆静电场轨道阱高分辨质谱联用技术(UPLC-Q-Exactive)结合中药成分高分辨质谱数据库(OTCML),对沙棘果茎叶的化学成分进行了快速鉴定与比较分析。色谱柱为HypersilGOLDaQ色谱柱(2.1mm×100mm,1.9μm),流动相为0.1%甲酸乙腈(A)-0.1%甲酸水(B)梯度洗脱,流速为0.3mL/min,高分辨质谱采用电喷雾离子源(ESI)正、负离子同时扫描模式。基于OTCML中所包含的1200余种中药化合物对照品的保留时间,分子离子、二级碎片离子谱图信息和文献数据进行结构鉴定。结果共鉴定出沙棘中102个功效成分,包括苷类18个,黄酮类21个,萜类18个,苯丙素类13个,生物碱类6个,挥发成分14个,多酚12个;其中有17个化合物是在沙棘中首次发现,对沙棘果茎叶比较表明这些功效成分在不同部位分布有明显差异。该方法为今后充分开发利用沙棘资源提供了基础数据,也为沙棘相关产品的质量控制和药效物质基础研究奠定了基础。     

基于线粒体细胞色素氧化酶I（COI）序列的猪和兔四个疥螨分离株的系统发育关系分析

为了探讨兔疥螨分离株和猪疥螨分离株的分类地位, 采用PCR技术首次扩增了分离自中国猪和兔的4个疥螨分离株的线粒体细胞色素氧化酶I（COI）基因, 并与GenBank中注册的14个国外疥螨分离株的同源基因进行了比较。序列分析结果显示: 扩增的4个疥螨株COI基因长度均为1 427 bp, 序列间无插入、缺失, A+T含量（73%）明显高于G+C含量（27%）, 碱基组成存在明显偏移。猪和兔的4个疥螨分离株间的COI基因同源性较高（99.1%～100.0%）, 它们与澳大利亚人疥螨株、国外动物疥螨株的同源性范围为98.4%～99.6%。在构建的NJ树中, 分离自中国猪和兔的4个疥螨分离株同澳大利亚人疥螨分离株、国外动物疥螨分离株亲缘关系较近。根据疥螨COI基因同源性分析和系统树构建结果, 我们认为分离自中国猪和兔的4个疥螨分离株与澳大利亚人疥螨分离株以及国外的动物疥螨分离株均应属于同一个种。     

聚丁二酸丁二醇酯降解菌株的筛选及其降解性能研究

从石油污染土壤中筛选到1株具有聚丁二酸丁二醇酯降解能力的菌株PBS1302,经菌体形态特征、菌落培养特征、生理生化鉴定和16S r DNA基因序列分析,初步鉴定为铜绿假单胞菌(Pseudomonas aeruginosa)。该菌株在培养温度37℃,培养基起始p H 6.8的条件下经6 d的培养,对聚丁二酸丁二醇酯薄膜的降解率可达36.9%。经电子显微镜观察,与降解前相比,聚丁二酸丁二醇酯薄膜表面变得粗糙,出现明显的蚀刻痕迹。     

The Hb A variant (beta73 Asp-->Leu) disrupts Hb S polymerization by a novel mechanism

Polymerization of a 1:1 mixture of hemoglobin S (Hb S) and the artificial mutant HbAbeta73Leu produces a dramatic morphological change in the polymer domains in 1.0 M phosphate buffer that are a characteristic feature of polymer formation. Instead of feathery domains with quasi 2-fold symmetry that characterize polymerization of Hb S and all previously known mixtures such as Hb A/S and Hb F/S mixtures, these domains are compact structures of quasi-spherical symmetry. Solubility of Hb S/Abeta73Leu mixtures was similar to that of Hb S/F mixtures. Kinetics of polymerization indicated that homogeneous nucleation rates of Hb S/Abeta73Leu mixtures were the same as those of Hb S/F mixtures, while exponential polymer growth (B) of Hb S/Abeta73Leu mixtures were about three times slower than those of Hb S/F mixtures. Differential interference contrast (DIC) image analysis also showed that fibers in the mixture appear to elongate between three and five times more slowly than in equivalent Hb S/F mixtures by direct measurements of exponential growth of mass of polymer in a domain. We propose that these results of Hb S/Abeta73Leu mixtures arise from a non-productive binding of the hybrid species of this mixture to the end of the growing polymer. This "cap" prohibits growth of polymers, but by nature is temporary, so that the net effect is a lowered growth rate of polymers. Such a cap is consistent with known features of the structure of the Hb S polymer. Domains would be more spherulitic because slower growth provides more opportunity for fiber bending to spread domains from their initial 2-fold symmetry. Moreover, since monomer depletion proceeds more slowly in this mixture, more homogeneous nucleation events occur, and the resulting gel has a far more granular character than normally seen in mixtures of non-polymerizing hemoglobins with Hb S. This mixture is likely to be less stiff than polymerized mixtures of other hybrids such as Hb S with HbF, potentially providing a novel approach to therapy.     

透明颤菌血红蛋白基因在解淀粉芽孢杆菌中的表达及其影响

将强启动子P43与透明颤菌血红蛋白基因（vgb）通过重叠延伸PCR进行融合,克隆到芽孢杆菌整合表达载体pDG1730中,重组表达载体pDG-P43vgb转化促生防病解淀粉芽孢杆菌FZB42,Wsetern-Blot和CO差光谱分析表明重组菌株FZB42-VHb表达了有活性的VHb蛋白,VHb的表达对重组菌株菌体的生长及抗菌脂肽的产生都有促进作用.在相同培养条件下,重组菌最大菌体密度比原始菌株提高了14.49 %,抗菌脂肽fengycin的产量提高了1.74倍,抗菌脂肽surfactin的产量提高了3.14倍.     

异常血红蛋白HbQueens及Hb广州—杭州的携氧功能及78—180K的穆斯堡尔谱测定

本实验通过对HbQueens[α_(34)B_(15)Leu→Arg]及Hb广州-杭州[α_(64)E_(13)Asp→Gly]的氧亲合力及它们在78-180K范围的穆斯堡尔谱测定,得到氧亲合力大小顺序为HbQueens>Hb广州-杭州>HbA,由穆斯堡尔谱结果的计算所得的结构参数表明中心Fe~(2+)与卟啉环平面的距离为HbQueens相似文献   

A Proteinase from Mung Bean Sprouts That Inactivaties Soybean Trypsin Inhibitor (in English)

By 30%-60% (NH4)2SO4 fractional precipitation, anion-exchange chromatography on DEAE-Sepharose CL-6B, gel filtration on Sephacryl S-200 and anion-exchange chromatography on Waters AP-1 column (ProteinPM-Pak DEAE 15HR), a proteinase which can inactivate soybean trypsin inhibitor (STI) was purified from mung bean (Vigna rabiata (L.) Wilczek) sprouts. Its molecular weight was estimated to be 29.8 kD by SDS-PAGE, and its Km and Vmax for STI were 769.2N-α-benzoyl-L-arginine ethyl ester BAEE/mL and 115.3 BAEE·mL-1·min-1 respectively. This proteinase was stable at temperatures lower than 50℃ and pH 6.5-8.5, and 90.91% STI activity of defatted soybean powder was inactivated by this preparation, with proteolytic activity 5 000 BAEE/mL at 50℃ and pH 8.0 in 4 h.     

紫外线诱变选育聚丁二酸丁二醇酯（PBS）高降解活性菌株

以PBS降解菌HJ03（Alternariasp．）为出发菌株,通过紫外诱变,透明圈初筛及PBS薄膜复筛,获得一株降解能力增强且对温度和pH耐受力均得到提高的突变株HJ10。与出发菌株相比,HJ10在培养初期气生菌丝少,而培养7d时菌落致密且生长速度较快。经过连续继代培养7代后发现,突变株的降解活力保持了良好的遗传稳定性,其降解率较出发菌株提高百分比达14．4％以上;在最适降解温度范围内（25℃～30℃）和不适宜降解的温度条件下突变菌株Ⅻ10的降解率均高于出发菌株;各pH条件下,突变株对PBS的降解能力明显优于出发菌株,尤其在pH5．0时降解率提高了22．80％。     

Resting-state coupling between HbO and Hb measured by fNIRS in autism spectrum disorder

To distinguish between children with autism spectrum disorder (ASD) and typically developing (TD) children, we have uncovered a new discriminative feature, hemoglobin coupling. Functional near-infrared spectroscopy (fNIRS) was used to record resting-state hemodynamic fluctuations in the bilateral temporal lobes in 25 children with ASD and 22 TD children, in which the coupling between low frequency oxygenated hemoglobin (HbO) and deoxygenated hemoglobin (Hb) fluctuations was evaluated by Pearson correlation coefficient. The results showed significantly weak coupling in children with ASD in both the left and right, and throughout the whole temporal cortex. To explain this observation, a simulation study was performed using a balloon model, in which we found four related parameters could impact the coupling. This study suggested that hemoglobin coupling might be applied as a new cerebral hemodynamic characteristic for ASD screening or diagnostics.     

Insect Trypsin Modulating Oostatic Factor (Neb-TMOF) and Its Analogs: Preliminary Structure/Biological Function Relationship Studies

Summary Neb-TMOF, the trypsin modulating oostatic factor of gray fleshflyNeobellieria bullata, is a hexapeptide with the following sequence: H-Asn-Pro-Thr-Asn-Leu-His-OH. It has been isolated from vitellogenic ovaries in 1994. TMOF, the newly discovered insect peptide, inhibits trypsin biosynthesis in the gut, lowers yolk polypeptide concentration in the hemolymph and strongly inhibits ecdysone biosynthesis by larval ring glands. It is interesting that this short non-protected peptide contains in its molecule two Asn residues at positions 1 and 4 and His at its C-terminus. To obtain information about the role of the His-6 and Asn-4 residues we synthesised two series of Neb-TMOF analogs, modified: (1) in position 6 byd-His (I), His(Bzl) (II) and Phe(p-X) derivatives, where X=NH₂ (III), NO₂ (IV), OEt (V) and OH (VI) and (2) in position 4 by such amino acid residues as Ser (VII), Thr (VIII), Gly (IX), Asp (X), Glu (XI) andd-Asn (XII). The influence of these peptides on trypsin biosynthesis inN. bullata was determinedin vivo. In preliminary investigations, we found that Neb-TMOF, [Phe(NH₂)⁶], and [Phe(NO₂)⁶]-Neb-TMOF inhibited trypsin biosynthesis, whereas [d-His)⁶]- and [d-His(Bzl)⁶]-Neb-TMOF were inactive. In further biological studies performedin vitro on heart ofTenebrio molitor were found that-TMOF and [Phe(p-NH₂)⁶]-Neb-TMOF showed weak cardioexcitatory activity, about 30% of the cardioexcitatory activity of proctolin, an insect neuromodulating peptide.     

Insect Trypsin Modulating Oostatic Factor (Neb-TMOF) and Its Analogs: Preliminary Structure/Biological Function Relationship Studies

Neb-TMOF, the trypsin modulating oostatic factor of gray fleshfly Neobellieria bullata, is a hexapeptide with the following sequence: H-Asn-Pro-Thr-Asn-Leu-His-OH. It has been isolated from vitellogenic ovaries in 1994. TMOF, the newly discovered insect peptide, inhibits trypsin biosynthesis in the gut, lowers yolk polypeptide concentration in the hemolymph and strongly inhibits ecdysone biosynthesis by larval ring glands. It is interesting that this short non-protected peptide contains in its molecule two Asn residues at positions 1 and 4 and His at its C-terminus. To obtain information about the role of the His-6 and Asn-4 residues we synthesised two series of Neb-TMOF analogs, modified: (1) in position 6 by D-His (I), His(Bzl) (II) and Phe(p-X) derivatives, where X = NH₂ (III), NO₂ (IV), OEt (V) and OH (VI) and (2) in position 4 by such amino acid residues as Ser (VII), Thr (VIII), Gly (IX), Asp (X), Glu (XI) and D-Asn (XII). The influence of these peptides on trypsin biosynthesis in N. bullata was determined in vivo. In preliminary investigations, we found that Neb-TMOF, [Phe(NH₂)⁶], and [Phe(NO₂)⁶]-Neb-TMOF inhibited trypsin biosynthesis, whereas [D-His)⁶]- and [D-His(Bzl)⁶]-Neb-TMOF were inactive. In further biological studies performed in vitro on heart of Tenebrio molitor we found that Neb-TMOF and [Phe(p-NH₂)⁶-Neb-TMOF showed weak cardioexcitatory activity, about 30% of the cardioexcitatory activity of proctolin, an insect neuromodulating peptide.