嗜热链球菌抗噬菌体菌株的选育

研究以酸奶生产菌株嗜热链球菌S2为出发菌株。采用自然选育的方法筛选出其抗噬菌体突变株。获得4株生产性能较好的抗噬菌株KS21、KS22、KS23、KS24,溶源性检测其为非溶源菌。对4株抗性菌株进行20次传代和酸奶发酵实验,结果表明抗性稳定,其中KS23发酵酸奶产酸达109°T,凝乳时间与出发菌株相当。细胞全蛋白SDS-PAGE电泳分析显示突变菌株与出发菌株在某些蛋白表达上有所差异,可能是抗性产生的原因。     

嗜热链球菌自发突变抗噬菌体菌株的选育及其特性

以酸奶发酵常用菌株嗜热链球菌为出发菌株,反复使用高密度噬菌体培养处理,初步获得9株抗噬菌体突变株。对抗性菌株进行遗传稳定性、溶源性、EOP检测,及抗性菌株正常条件下发酵性能测定,综合各方面特性,筛选出遗传稳定,生长、发酵性能优于野生菌株的抗性菌株,对优于野生株的抗性菌株与敏感菌株一起在较高浓度噬菌体的环境中进一步发酵,测定其对噬菌体的抵抗能力、产酸能力、发酵后酸奶的黏度、硬度、乳清析出、口感及后酸化程度。结果表明:9株抗性菌株有6株遗传性比较稳定,其均为非溶源性菌,其中有3株生长能力、产酸能力、发酵乳黏度、硬度、接近于野生菌。这3株抗性菌株在高噬菌体浓度下发酵,有2株菌生长产酸正常,黏度、硬度、口感较好,凝乳时间较短、乳清析出较少并且后酸化程度相对较弱,适宜工业生产。      

嗜热链球菌抗噬菌体菌株的选育及其发酵特性

为了筛选到适合酸奶生产的抗噬菌体菌株,采用双层平板法及自然选育的方法,从酸奶异常发酵液中分离到3种不同的噬菌体,并获得了4株具有抗这些噬菌体能力的非溶源性抗性菌株St1、St2、St3、St4,对这4株抗性菌株进行20次传代和酸奶发酵实验,结果表明其抗性稳定,其中St3发酵酸奶产酸可达110°T,凝乳时间与出发菌株基本相当,在高噬菌体浓度下发酵,产酸正常,黏度、硬度、口感较好,凝乳时间较短、乳清析出较少并且富集培养后菌体浓度可达1010CFU/mL,适宜于工业生产。     

单亲灭活保加利亚乳杆菌与嗜热链球菌抗噬菌体菌株融合子的选育

为获得具有抗噬菌体功能且发酵性能优良的乳酸菌酸奶生产菌株,将生产用保加利亚乳杆菌制备原生质体后高温灭活,灭活后的原生质体与嗜热链球菌抗噬菌体菌株的原生质体通过PEG6000诱导进行融合,并对融合子性能进行研究。结果从20株融合子中挑选出3株具有噬菌体抗性且发酵性能优良的乳酸菌融合子。融合条件为:以PEG6000（浓度为400 g/L,添加0.01 mol/L Ca Cl2、0.02 mol/L Mg Cl2）为促融剂,40℃融合2 min。此条件下,融合率可达1.85×10-6。获得的融合子在凝乳、产酸、产粘性物质、产香气成分及抗噬菌体方面性能优越,适合于酸奶生产。      

保加利亚乳杆菌与嗜热链球菌抗噬菌体菌株的原生质体制备及融合

为获得具有抗噬菌体功能且发酵性能优良的乳酸菌融合子,采用单亲灭活及正交分析方法,研究了保加利亚乳杆菌与嗜热链球菌抗噬菌体菌株的原生质体制备及融合条件。结果表明：保加利亚乳杆菌原生质体制备的最适条件是以磷酸盐缓冲液和甘露醇制作的高渗溶液为原生质体稳定剂,1.0 mg/mL的溶菌酶36 ℃处理30 min,原生质体的形成率为（89.02±2.31）%,再生率为（4.62±0.22）%。嗜热链球菌抗噬菌体菌株原生质体制备的最适条件是以Tris-HCl和蔗糖制作的高渗溶液为原生质体稳定剂,0.1 mg/mL的溶菌酶42 ℃处理30 min,原生质体的形成率为（99.15±0.23）%,再生率为（5.79±0.17）%。单亲灭活保加利亚乳杆菌与嗜热链球菌抗噬菌体菌株原生质体融合的最适条件为聚乙二醇6000（质量浓度为400 g/L,添加0.01 mol/L CaCl2、0.02 mol/L MgCl2）40 ℃促融2 min,融合率可达（1.85±0.12）×10－6。所得融合子各项性能优良,适合于酸奶生产。     

嗜热链球菌噬菌体对酸奶质量的影响

研究嗜热链球菌噬菌体对酸奶直投式发酵剂(direct vat set,DVS)发酵产酸、嗜热链球菌与保加利亚乳杆菌比例、酸奶黏度、口感、后酸化、乳清析出等方面的影响。结果表明：嗜热链球菌噬菌体可影响酸奶发酵剂发酵产酸、明显改变酸奶嗜热链球菌与保加利亚乳杆菌比例,导致产品黏度降低、口感变差、乳清析出及后酸化严重,嗜热链球菌噬菌体对DVS 生产酸奶有着严重的危害。     

嗜热链球菌噬菌体的分离纯化与鉴定

噬菌体颗粒的分离纯化是研究噬菌体生物学特性、基因分析的基础工作。本研究采用聚乙二醇浓缩噬菌体增殖液、缓冲液透析去除聚乙二醇、氯化铯密度梯度离心纯化,得到了纯度较高的嗜热链球菌噬菌体样品。多重PCR鉴定和SDS-PAGE蛋白组成分析结果表明,该嗜热链球菌噬菌体的包装机制为pac-型。      

嗜热链球菌噬菌体的分离纯化与鉴定

噬菌体颗粒的分离纯化是研究噬菌体生物学特性、基因分析的基础工作。本研究采用聚乙二醇浓缩噬菌体增殖液、缓冲液透析去除聚乙二醇、氯化铯密度梯度离心纯化,得到了纯度较高的嗜热链球菌噬菌体样品。多重PCR鉴定和SDS-PAGE蛋白组成分析结果表明,该嗜热链球菌噬菌体的包装机制为pac-型。     

抗噬菌体保加利亚乳杆菌突变菌株L.actobacillus burglarious-23UN的选育

在酸奶发酵生产过程中,易遭受噬菌体的感染,使生产不能进行.以高密度噬菌体培养处理保加利亚乳杆菌菌株,筛选出1株噬菌体抗性菌株L-13,但其产酸、产黏、产香能力不如原菌株.通过照射距离20 cm照射时间60 s的紫外线诱变和NTG振荡70min的复合诱变,得到LUN-23菌株,其产酸力较敏感菌株提高了2.6%,产粘性提高了2.2%,产乙醛能力提高了1.1%,产丁二酮能力提高了3.8%,且具有遗传稳定性和抗109pfu/mL噬菌体浓度的能力.     

超高压对嗜热链球菌噬菌体灭活效果的研究

为研究超高压(HHP)对嗜热链球菌噬菌体的灭活效果,以3株嗜热链球菌噬菌体为对象,在不同压力、处理时间及初始菌数条件下进行超高压处理,并与传统热处理进行比较;通过观察处理前、后噬菌体结构的变化,探明超高压灭活机理。试验结果表明,压力400MPa下无明显灭活效果;压力600MPa处理10~20min,可将3株噬菌体全部杀灭。噬菌体ALQ13.2和DT1随初始菌数的下降,灭活效果逐渐增强;Abc2则无显著性差异。72℃、15s的传统热处理对噬菌体无明显灭活作用,而时间延长至5min,可将大部分噬菌体杀灭。3株噬菌体的耐压性与耐热性无相关性,其中ALQ13.2最耐压,DT1最耐热。电镜结果显示超高压处理前、后噬菌体的结构发生多种变化。     

紫外诱变选育高产3-羟基丁酮枯草芽孢杆菌

选育高产3-羟基丁酮的枯草芽孢杆菌新菌株。采用紫外诱变(15W,25cm),对一株产3-羟基丁酮枯草芽孢杆菌YD-1进行紫外诱变处理,并对菌株的遗传稳定性进行测定。筛选获得4株高产菌MB-2、MB-6、MB-10和MB-11,经过20代传代培养后MB-2的遗传稳定性最好,其遗传物质经RAPD分析,与出发菌株YD-1相比有较大差异。该菌遗传性状稳定,是一株高产3-羟基丁酮的新菌株。     

紫外诱变选育高产蛋白酶枯草芽孢杆菌

紫外线辐射是目前最为广泛的物理诱变因子之一,其引起菌体DNA结构变化的形式很多,如DNA链的断裂、碱基的破坏等,但最主要的作用是使同链DNA的相邻嘧啶间形成胸腺嘧啶二聚体,阻碍碱基间的正常配对,从而引起微生物突变或死亡.以实验室保存的枯草芽孢杆菌为出发菌株,利用紫外诱变的方法,以获取可高效发酵生产蛋白酶的突变株.通过大量筛选及连续传代实验,确定了一株突变性状能稳定遗传的突变株,其蛋白酶产量较出发菌株有大幅度提高.     

烟草抗PVY育种材料的筛选与应用

马铃薯Ｙ病毒对东北烟区烟草生产危害日趋严重。通过病圃发病情况调查和温室接种鉴定,初步筛选出ＣＶ９１、８７４１４、８０２１三份常规抗马铃薯Ｙ病毒材料,其中ＣＶ９１在田间调查和温室接种鉴定中表现为高抗马铃薯Ｙ病毒;转基因抗马铃薯Ｙ病毒材料８５－２、８６－１,８７－２,８９－１、９４－１、９５－１、９８－１温室接种鉴定结果为对ＰＶＹ免疫,利用ＣＶ９１育成了中抗ＰＶＹ的新品系９１１１－２１。     

一株多重耐药副溶血性弧菌的耐药分子机制研究

目的从养殖半滑舌鳎肠道分离到一株副溶血性弧菌Vp1107,对该菌株进行耐药性及耐药分子机制研究。方法通过K-B法进行药敏试验,PCR扩增及测序法对耐药基因及整合子进行检测与分析。结果 Vp1107菌株耐受氨苄西林、阿莫西林、头孢唑啉、四环素、土霉素和氯霉素,对头孢呋辛钠、链霉素、卡那霉素、萘啶酸、环丙沙星中度敏感,其多重耐药系数为0.33。菌株携带blaTEM、strA、strB、tetB、tetM、catⅢ、intI1耐药相关基因,1类整合子不含基因盒,gyrA和parC基因的喹诺酮类耐药决定区未发生点突变。结论副溶血性弧菌Vp1107耐药程度较为严重,其耐药性主要由耐药基因编码的耐药性酶类和外排泵作用引起,提示应加强对副溶血性弧菌耐药性的监控及渔用药物的管理。     

Studies on resistance to insecticides in Tribolium castaneum (Herbst)—III. Selection of a strain resistant to lindane and its biological characteristics

A strain of Tribolium castaneum (Herbst) resistant to lindane was selected from a heterogeneous parent stock by continuous breeding in wheat flour treated with lindane. Starting with 20 ppm in the first, the concentration of lindane in each subsequent generation was increased till a concentration of 600 ppm was reached in the ninth generation. A tenth generation was reared in 600 ppm lindane and the adults which emerged were used for further investigations.

The resistance was estimated by three testing methods, namely, treatment of the rearing medium, film exposure, and direct spray. The level of resistance was assessed in comparison with (i) the parental strain, and (ii) the susceptible strain developed when the parental stock was reared for ten generations without any insecticidal treatment. The lindane-resistant strain when tested by rearing medium, film exposure and direct spray methods was found to be 54·8, >87, and 13·05 times as resistant as the parental, and 90·2, >86, and 12·48 times as resistant as the susceptible strain respectively.

The resistant strain had a lower fecundity than the susceptible strain, but did not show any change in the duration of the preimaginal stages when reared in untreated medium. However, there was a prolongation of the larval period in the rearing medium treated with lindane.     

Creating a Saccharomyces cerevisiae haploid strain having 21 chromosomes

Chromosome engineering techniques that can manipulate a large segment of chromosomal DNA are useful not only for studying the organization of eukaryotic genomes but also for the improvement of industrially important strains. Toward the development of techniques that can efficiently manipulate a large segment of chromosome, we have previously reported a one-step chromosome splitting technique in a haploid Saccharomyces cerevisiae cell, with which we could successfully split yeast chromosome 11, XIII, or XI into two halves to create a haploid strain having 17 chromosomes. We have now constructed chromosome splitting vectors bearing ADE2, HIS3, LEU2, or TRP1 marker, and by using these vectors, we could successively split yeast chromosomes to create a novel yeast haploid strain having up to 21 chromosomes. The specific growth rates of yeast strains carrying more than 16 chromosomes up to 21 did not differ significantly, suggesting that yeast cells can harbor more chromosomes than they do in their natural state, that is, 16 chromosomes, without serious effects on their growth.     

Sequencing of plasmids from a multi-antimicrobial resistant Salmonella enterica serovar Dublin strain

Salmonella enterica serovar Dublin (S. Dublin) is a host-adapted serotype whose primary host is cattle, which can serve as a potential reservoir for human infections. S. Dublin remains one of the leading causes of severe invasive infections and deaths associated with salmonellosis. Because of their propensity to cause severe infection, antimicrobial therapy is often required, thus antimicrobial resistance is an important concern. Plasmids play a key role in facilitating drug resistance in these pathogens. This study reports the results of DNA sequencing and sequence analysis of plasmids from a highly multidrug resistant strain (resistant to 11/15 drugs tested) of S. Dublin that originated from cattle. The strain was found to contain four plasmids of approximately 8, 77, 89, and 174 kb. The 174 kb plasmid is an incompatibility group (Inc) A/C plasmid containing genes associated with resistance to at least 9 different antimicrobials, as well as disinfectants and metals. The 88.5 kb plasmid is an IncFIB plasmid containing genes associated with resistance to at least 3 antimicrobial agents and mercurial compounds. The 77 kb plasmid is a S. Dublin virulence plasmid containing multiple virulence-associated genes and the 7.9 kb plasmid encodes mobilization and replication genes. Overall, sequencing identified multiple plasmids containing antimicrobial resistance and virulence genes. The resistance genes identified correlated to the observed resistance phenotype, further indicating the importance of plasmids in antimicrobial resistance in many Salmonella.     

Medical and biological assessment of genetically modified corn line MON 810 resistant to European corn borer and line GA 21 resistant to glyphosate: a chemical study

Chemical analysis of genetically modified corns MON 810 resistance to European corn borer and GA 21 tolerance to glyphosphate was performed. Results of these studies showed that there is no difference between genetically modified and conventional corn products.     

紫外诱变和丁醇驯化复合选育高产丁醇菌株

本研究以Clostridium beijerinckii NCIMB 8052为出发菌株,利用紫外诱变和高丁醇环境驯化相结合的方法复合选育,最终获得一株耐丁醇的高产突变株,命名为Clostridium beijerinckii ZL01.与出发菌株8052相比,ZL01对丁醇初始浓度的耐受能力从10g/L提高到11g/L.5L发酵罐的分批发酵结果表明,丁醇的产量从10.34g/L增加为15.01g/L,提高45.16％;总溶剂从12.87g/L增加为19.55g/L,提高49.01％;发酵周期缩短4h,发酵强度从0.27g/(L·h)提高为0.44g/(L·h),遗传稳定性实验表明,该菌株连续传代20次,溶剂产量稳定,菌株无明显退化.     

抗赤星病烟草的防卫基因的表达与基因组DNA结构的变化

用赤星病菌（Ａｌｔｅｒｎａｒｉａ ａｌｔｅｍａｔａ）毒性菌株产生的ＡＴ毒素诱导烟草,可使寄主获得对赤星病８０．４％的系统抗性,比病菌弱毒株ＴＢＡ１６孢子诱导的效果高约２０％。烟草品种ＮＣ８９心叶叶碟在含ＡＴ毒素６０％和７０％的ＭＳ培养基上经２轮胁迫筛选和植株再生,得到抗病品系ＮＣ８９－ＴＴ。用５种防卫基因的ｃＤＮＡ探针检测了不诱导、受ＡＴ毒素诱导、受病菌弱毒株激发子诱导的ＮＣ８９和诱导与不诱导的ＮＣ８９－ＴＴ第４代植株５种基因的转录活性,结果表明,苯丙氨酸氨裂解酶（ＰＡＬ）基因在ＮＣ８９中不能组成型表达,可被ＡＴ毒素诱导激活,但不受激发子诱导;病程相关蛋白（ＰＲ蛋白）ＰＲ－ｌａ、查尔酮合成酶（ＣＨＳ）、脂氧合酶（ＬＯＸ）基因在ＮＣ８９中表现为诱导转录或诱导后转录活性增强;这４种基因都在ＮＣ８９－ＴＴ中组成型表达