非编码的mRNA

非编码的mRNA是近年发现的一类不含典型ORF的mRNA.目前已发现或克隆的这类基因主要有：H19基因,XIST基因,XLSIRT基因,His-1基因,bic基因,rox1和rox2基因等.它们与胚胎发育,肿瘤发生及X染色体失活密切相关.     

Osmoregulation in the model organismEscherichia coli: genes governing the synthesis of glycine betaine and trehalose and their use in metabolic engineering of stress tolerance

Glycine betaine is known to be the preferred osmoprotectant in many bacteria, and glycine betaine accumulation has also been correlated with increased cold tolerance. Trehalose is often a minor osmoprotectant in bacteria and it is a major determinant for desiccation tolerance in many so-called anhydrobiotic organisms such as baker's yeast(Saccharomyces cerevisiae). Escherichia coli has two pathways for synthesis of these protective molecules; i.e., a two-step conversion of UDP-glucose and glucose-6-phosphate to trehalose and a two-step oxidation of externally-supplied choline to glycine betaine. The genes governing the choline-to-glycine betaine pathway have been studied inE. coli and several other bacteria and higher plants. The genes governing UDP-glucose-dependent trehalose synthesis have been studied inE. coli andS. cerevisiae. Because of their well-documented function in stress protection, glycine betaine and trehalose have been identified as targets for metabolic engineering of stress tolerance. Examples of this experimental approach include the expression of theE. coli betA andArthrobacter globiformis codA genes for glycine betaine synthesis in plants and distantly related bacteria, and the expression of theE. coli otsA and yeastTPS1 genes for trehalose synthesis in plants. The published data show that glycine betaine synthesis protects transgenic plants and phototrophic bacteria against stress caused by salt and cold. Trehalose synthesis has been reported to confer increased drought tolerance in transgenic plants, but it causes negative side effects which is of concern. Thus, the much-used model organismE. coli has now become a gene resource for metabolic engineering of stress tolerance.     

地黄C3H基因的克隆及生物信息学分析

香豆酸-3-羟化酶属于植物中最大的蛋白酶细胞色素P450家族之一,在植物生命活动中发挥着重要作用。为了解地黄香豆酸-3-羟化酶基因RgC3H合成毛蕊花糖苷的功能,该研究基于地黄代谢组学分析获得KEGG途径中的C3H,采用多重比对在NCBI中获得同源基因的一个保守序列,并基于该保守序列和地黄SRA数据库,采用电子克隆和RT-PCR克隆技术获得地黄C3H基因全长CDS(RgC3H),对其进行生物信息学分析。结果表明:RgC3H基因全长为1 530 bp,且编码一个含509个氨基酸、分子量为57.91 kD、无信号肽的蛋白质; 基于氨基酸序列的结构分析显示,RgC3H有一个保守区域-P450结构域; 系统进化分析结果显示,RgC3H与芝麻和猴面花的C3H基因具有很高的同源性。上述结果为进一步研究RgC3H基因在地黄毛蕊花糖苷生物合成途径中的作用奠定了基础。     

The Drosophila Tumorous lethal hematopoietic oncogene is a dominant mutation in the hopscotch locus

The Drosophila Tumorous-lethal (Tum-l) mutation acts as an activated oncogene, causing hematopoietic neoplasms, overproliferation, and premature differentiation. Tum-l is a dominant mutation in the hopscotch (hop) locus, which is required for cell division and for proper embryonic segmentation. The Tum-l temperature-sensitive period for melanotic tumor formation includes most of larval and pupal development.     

Cloning, sequencing and expression of a recA gene from Amycolatopsis mediterranei

The 3.5 kb nucleotide fragment, including the recA gene and its downstream recX-like gene, has been isolated from a genomic library by dot-blot hybridization with the Mycobacterium smegmatis recA gene. The recA gene, consisting of 1047 base pairs (bp), encodes a polypeptide of 348 amino acids while the recX-like gene, consisting of 450 bp, encodes a shorter polypeptide of 149 amino acids. Both the deduced amino acid sequences of recA and recX resemble those of the recA and recX genes from other bacteria. The cloned Amycolatopsis mediterranei U32 recA gene conferred partial resistance to ethyl methane sulfonate when expressed in E. coli with the lacZ promoter.     

不同代次牛成肌细胞的诱导分化及相关基因的表达分析

利用初生荷斯坦牛的成肌细胞,在不同代次以含体积分数为2%马血清的DMEM进行诱导分化,在之后0、2、4、6、8、10 d观察细胞的形态变化并收集细胞提取总RNA,检测成肌相关基因的表达情况,为进一步研究牛肌肉发生过程及相关基因的表达调控提供依据。同时利用实时荧光定量PCR分别检测肌性相关基因MyoD(生肌决定因子)、MyoG(肌细胞生成素)以及非肌性相关基因A-FABP(脂肪细胞型脂肪酸结合蛋白)表达水平的变化。研究表明：①各代细胞在诱导培养4 d后开始有肌管形成,其后越来越多,8 d时达高峰。②成肌细胞向成熟肌细胞分化过程中,MyoD、MyoG、A-FABP基因都呈先上升然后下降的趋势。③高代次成肌细胞比低代次细胞增殖慢,而且在诱导分化后,肌管的数目明显变少; MyoD、MyoG、A-FABP随着代次的升高而下降。故推断MyoD、MyoG以及A-FABP基因会在成肌分化开始后的不同阶段被激活,从而发挥不同的调控作用,而且随着传代次数的增加成肌细胞的分化能力减弱。     

Anaerobic bacteria and antibiotics: What kind of unexpected resistance could I find in my laboratory tomorrow?

The purpose of this article is to set out some important considerations on the main emerging antibiotic resistance patterns among anaerobic bacteria. The first point concerns the Bacteroides fragilis group and its resistance to the combination of β-lactam+β-lactamase inhibitor. When there is overproduction of cephalosporinase, it results in increased resistance to the β-lactams while maintaining susceptibility to β-lactams/β-lactamase inhibitor combinations. However, if another resistance mechanism is added, such as a loss of porin, resistances to β-lactam+β-lactamase inhibitor combinations may occur. The second point is resistance to metronidazole occurring due to nim genes. PCR detection of nim genes alone is not sufficient for predicting resistance to metronidazole; actual MIC determinations are required. Therefore, it can be assumed that other resistance mechanisms can also be involved. Although metronidazole resistance remains rare for the B. fragilis group, it has nevertheless been detected worldwide and also been observed spreading to other species. In some cases where there is only a decreased susceptibility, clinical failures may occur. The last point concerns resistance of Clostridium species to glycopeptides and lipopeptides. Low levels of resistance have been detected with these antibiotics. Van genes have been detected not only in clostridia but also in other species. In conclusion, antibiotic resistance involves different mechanisms and affects many anaerobic species and is spreading worldwide. This demonstrates the need to continue with antibiotic resistance testing and surveys in anaerobic bacteria.     

酮型广藿香MVA途径基因表达分析及与主要成分合成相关性研究

广藿香药材以广藿香酮含量较高的酮型广藿香为最优质。而广藿香酮为一种萜类成分,其生物合成途径尚未明确。MVA(甲羟戊酸)途径是萜类化合物生物合成的重要途径。为了分析MVA途经基因表达与化学成分的相关性从而获得促进广藿香酮合成的潜在基因,该文以2种酮型广藿香栽培品种(石牌广藿香、高要广藿香)为材料,通过实时定量PCR分析基因表达和主要成分含量测定,并研究了供试材料不同时期的茎、叶中与甲羟戊酸代谢途径相关的HMGR、MK、MDD基因表达及化学成分。结果表明：(1)HMGR基因在石牌广藿香嫩叶中表达更明显;MK基因在石牌广藿香和高要广藿香中表达模式相似,主要在老茎中表达;MDD基因在石牌广藿香叶中比高要广藿香表达量更高,在两种广藿香的茎中表达模式相似。(2)同属于酮型广藿香,石牌广藿香与高要广藿香的化学成分相似,老叶广藿香醇含量最高,老茎的广藿香酮含量更高。(3)MDD和MK基因与广藿香酮的合成正相关。综上结果所述,酮型广藿香两个栽培种MVA途径的基因表达模式相似,MDD和MK基因可能为酮型广藿香萜类代谢途径的关键基因。     

红曲霉繁殖相关brlM基因鉴定及功能分析

brlA作为曲霉分生孢子形成的中心调控路径中最上游的转录因子,能够诱导下游特异基因的表达调控分生孢子的产生,brlA缺失将导致曲霉无法产生分生孢子,同时生长代谢发生改变,但对不同菌种的影响有显著差异。【目的】探究brlA同源基因brlM在红曲霉中的基因功能,探索红曲霉繁殖调控机制。【方法】从紫色红曲霉Mp-21菌株中克隆了brlA同源基因brlM。利用同源重组原理,用农杆菌介导转化法构建了brlM基因缺失突变株(△brlM),分析△brlM和野生菌株Mp-21在菌落表型、显微结构、生长速率和次生代谢产物等方面的差异,明确brlM基因在红曲霉中的主要功能。【结果】在形态水平上,brlM基因缺失菌株导致菌丝生长更加旺盛,赋予菌落更加蓬松的表型;显微观察发现△brlM失去了有性繁殖产生闭囊壳的能力,但却提升了无性繁殖产生分生孢子的能力;同时代谢产物红曲色素、莫纳可林K和桔霉素产量显著下降。【结论】红曲霉brlM基因的功能与曲霉属中的brlA并不相同,brlM基因在红曲霉有性繁殖中的作用不可替代。本研究的结果对进一步探索丝状真菌繁殖调控机制提供了新的思路。     

Phylogenetic Studies of <Emphasis Type="Italic">Gordonia</Emphasis> Species Based on <Emphasis Type="Italic">gyrB</Emphasis> and <Emphasis Type="Italic">secA1</Emphasis> Gene Analyses

Phylogenetic relations within the genus Gordonia were analyzed using partial gyrB and secA1 gene sequences of 23 type species in comparison with those of 16S rRNA gene. The gyrB and secA1 phylogenies showed agreement with that constructed using 16S rRNA gene sequences. The degrees of divergence of the gyrB and secA1 genes were approximately 3.4 and 1.7 times greater, respectively, than that of 16S rRNA gene. The gyrB gene showed more discriminatory power than either the secA1 or 16S rRNA gene, facilitating clear differentiation of any two Gordonia species using gyrB gene analysis. Our data indicate that gyrB and secA1 gene sequences are useful as markers for phylogenetic study and identification at the species level of the genus Gordonia.     

Cis effect oflacZ sequences in transgenic mice

Transgenic mice carrying the 3-hydroxy-3-methylglutarylCoA reductase (HMG) promoter driving theEscherichia coli -galactosidase (lacZ) gene did not display the expected ubiquitous and constitutive expression inHMG-lacZ transgenic mice. The same promoter is however able to drive ubiquitous expression of the chloramphenicol acetyltransferase (cat) gene. Two lines of doubleHMG-lacZ andHMG-cat transgenic mice were obtained in which the two constructs were integrated at the same genomic sites. These mice expressed both reporter genes, but exclusively in the testes. These results suggest that thelacZ sequence might interfere negatively with the expression of the adjacentHMG-cat transgene.     

A new genetically isolated population of theSaccharomyces sensu stricto complex from Brazil

Genetic and karyotypic studies of someSaccharomyces sensu stricto yeasts from Brazil revealed a genetically isolated population which apparently represents a new sibling species ofS. cerevisiae.     

Stable chloroplast transformation in cabbage (<Emphasis Type="Italic">Brassica oleracea </Emphasis>L. var. <Emphasis Type="Italic">capitata </Emphasis>L.) by particle bombardment

The objectives of this research were first to isolate plastid gene sequences from cabbage (Brassica oleracea L. var. capitata L.), and to establish the chloroplast transformation technology of Brassica. A universal transformation vector (pASCC201) for Brassica chloroplast was constructed with trnV–rrn16S (left) and trnI–trnA–rrn23S (right) of the IR_A region as a recombination site for the transformed gene. In transforming plasmid pASCC201, a chimeric aadA gene was cloned between the rrn16S and rrn23S plastid gene borders. Expression of aadA confers resistance to spectinomycin and streptomycin antibiotics. The uidA gene was also inserted into the pASCC201 and transferred into the leaf cells of cabbage via particle gun mediated transformation. Regenerated plantlets were selected by 200 mg/l spectinomycin and streptomycin. After antibiotic selection, the regeneration percentage of the two cabbage cultivars was about 2.7–3.3%. The results of PCR testing and Southern blot analysis confirmed that the uidA and aadA genes were present in the chloroplast genome via homologously recombined. Northern blot hybridizations, immunoblotting and GUS histochemical assays indicated that the uidA gene were stable integrated into the chloroplast genome. Foreign protein was accumulated at 3.2–5.2% of the total soluble protein in transgenic mature leaves. These results suggest that the expression of a variety of foreign genes in the chloroplast genome will be a powerful tool for use in future studies.     

The Role of GIGANTEA Gene in Mediating the Oxidative Stress Response and in Arabidopsis

The Arabidopsis GIGANTEA (GI) gene has been shown to be involved in the regulation of the oxidative stress response; however, little is known about the mechanism by which GI gene regulates the oxidative stress response. We show here that enhanced tolerance of the gi-3 mutant to oxidative stress is associated, at least in part, with constitutive activation of superoxide dismutase (SOD) and ascorbate peroxidase (APX) genes. The gi-3 plants were more tolerant to parquart (PQ) or hydrogen peroxide (H₂O₂)-mediated oxidative stress than wild-type plants. Analyses of concentrations of endogenous H₂O₂ and superoxide anion radicals as well as lipid peroxidation revealed that enhanced tolerance of gi-3 plants to oxidative stress was not due to defects in the uptake of PQ or the sequestration of PQ from its site of action, and that the gi-3 mutation alleviated oxidative damage of plant cells from PQ stress. Moreover, the gi-3 mutant showed constitutive activation of cytosolic Cu/ZnSOD and plastidic FeSOD as well as cytosolic APX1 and stromal APX genes, which at least in part contributed to constitutive increases in activities of anti-oxidative enzymes SOD and APX, respectively. To our knowledge, we demonstrate, for the first time, that GI gene regulates the oxidative stress response, at least in part, through modulation of SOD and APX genes.     

Umchs5,a Gene Coding for a Class IV Chitin Synthase inUstilago maydis

A fragment corresponding to a conserved region of a fifth gene coding for chitin synthase in the plant pathogenic fungusUstilago maydiswas amplified by means of the polymerase chain reaction (PCR). The amplified fragment was utilized as a probe for the identification of the whole gene in a genomic library of the fungus. The predicted gene product ofUmchs5has highest similarity with class IV chitin synthases encoded by theCHS3genes fromSaccharomyces cerevisiaeandCandida albicans, chs-4fromNeurospora crassa,andchsEfromAspergillus nidulans. Umchs5null mutants were constructed by substitution of most of the coding sequence with the hygromycin B resistance cassette. Mutants displayed significant reduction in growth rate, chitin content, and chitin synthase activity, specially in the mycelial form. Virulence to corn plantules was also reduced in the mutants. PCR was also used to obtain a fragment of a sixth chitin synthase,Umchs6.It is suggested that multigenic control of chitin synthesis inU. maydisoperates as a protection mechanism for fungal viability in which the loss of one activity is partially compensated by the remaining enzymes.