基于16S rRNA基因和细菌基因组间重复序列的DNA指纹图谱技术对肝硬化大鼠肝移植后肠道菌群多样性的研究

目的应用PCR-DGGE和rep-PCR技术对肝硬化大鼠肝移植后肠道菌群多样性进行研究,探讨肠道菌群多样性的变化,并比较这两种方法在菌群分析中的作用。方法首先建立CCL4诱导肝硬化大鼠的肝移植模型,收取对照组、肝硬化成模时、肝移植后7 d和肝移植后30 d的大鼠粪便,提取细菌基因组DNA,采用PCR-DGGE和rep-PCR[BOX-PCR,ERIC-PCR,ERIC2-PCR,(GTG)5-PCR,REP-PCR]进行DNA指纹图谱分析。结果PCR-DGGE可明确将正常大鼠、肝硬化大鼠、肝移植大鼠分为3个簇,并显示出肝硬化、肝移植大鼠肠道菌群多样性明显增多。rep-PCR技术也可将各组分开,其中ERIC-PCR、ERIC2-PCR及REP-PCR三者扩增条带各组差异有显著性,鉴别效果更好。结论应用基于16S rRNA基因和细菌基因组间重复序列的指纹图谱技术对肝硬化大鼠肝移植后肠道微生态的研究具有重要价值,可对临床肝硬化、肝移植患者肠道微生态制剂的使用起到指导作用。     

银叶竹芋的组织培养和快速繁殖

1植物名称银叶竹芋(Ctenanthe setosa‘Greystar’)。2材料类别当年生茎节。3培养条件芽诱导培养基:(1)MS+BA1mg·L-1(单位下同)+NAA0.1;(2)继代增殖培养基MS+BA6+NAA0.05;(3)生根培养基MS+NAA0.5。以上培养基均附加3%蔗糖(生根培养基中为2%)、0.6%琼脂。pH5.8,培养温度(26±1)℃,每天光照12h,光强40μmol·m-2·s-1。4生长与分化情况4.1无菌材料的获得取当年生茎节,除去根状物,自来水冲洗表面20min。将茎节切成约2cm的小段,每段一芽,在超净工作台上,用75%的酒精浸泡30s,再置于0.1%升汞溶液(加1～2滴吐温20)中消毒10min,然后用无…     

Heteroplasmy levels of mtDNA1555A>G mutation is positively associated with diverse phenotypes and mutation transmission in a Chinese family

The mtDNA 1555A>G mutation was considered to be one of the most common causes of aminoglycoside-induced and non-syndromic hearing loss. However, this mutation was always found in homoplasmy with high phenotypic heterogeneity. Recently this mutation in heteroplasmy has been reported in several studies. In the present study, we have collected a large Chinese family harboring heteroplasmic mtDNA 1555A>G mutation with diverse clinical phenotypes. To investigate the relationship between the mutation load and the severity of hearing loss under Eastern Asian background, we performed clinical, molecular, genetic and phylogenic analysis. This pedigree was characterized by coexistence of eight subjects with homoplasmic mutation and ten subjects with various degrees of heteroplasmy, and the results suggested that there was a strong correlation between the mutation load and the severity/age-onset of hearing loss (r=0.758, p<0.001). We noticed that the mutation level of offspring was associated with their mothers' in this pedigree, which indicated that maybe exist a regular pattern during the process of the heteroplasmic transmission. In addition, analysis of the complete mtDNA genome of this family revealed that it belonged to Eastern Asian haplogroup B4C1. In addition, a rare homoplasmic mtDNA 9128T>C variant was identified, it located at a strictly conserved site of mtDNA ATP6 gene.     

Improved curdlan fermentation process based on optimization of dissolved oxygen combined with pH control and metabolic characterization of <Emphasis Type="Italic">Agrobacterium</Emphasis> sp. ATCC 31749

A significant problem in scale-down cultures, rarely studied for metabolic characterization and curdlan-producing Agrobacterium sp. ATCC 31749, is the presence of dissolved oxygen (DO) gradients combined with pH control. Constant DO, between 5% and 75%, was maintained during batch fermentations by manipulating the agitation with PID system. Fermentation, metabolic and kinetic characterization studies were conducted in a scale-down system. The curdlan yield, intracellular nucleotide levels and glucose conversion efficiency into curdlan were significantly affected by DO concentrations. The optimum DO concentrations for curdlan production were 45–60%. The average curdlan yield, curdlan productivity and glucose conversion efficiency into curdlan were enhanced by 80%, 66% and 32%, respectively, compared to that at 15% DO. No apparent difference in the gel strength of the resulting curdlan was detected. The comparison of curdlan biosynthesis and cellular nucleotide levels showed that curdlan production had positive relationship with intracellular levels of UTP, ADP, AMP, NAD⁺, NADH and UDP-glucose. The curdlan productivity under 45% DO and 60% DO was different during 20–50 h. However, after 60 h curdlan productivity of both conditions was similar. On that basis, a simple and reproducible two-stage DO control process for curdlan production was developed. Curdlan production yield reached 42.8 g/l, an increase of 30% compared to that of the single agitation speed control process.     

NtrC-dependent regulatory network for curdlan biosynthesis in response to nitrogen limitation in Agrobacterium sp. ATCC 31749

The mechanism of nitrogen signal regulating curdlan biosynthesis in Agrobacterium sp. ATCC 31749 was investigated. Under nitrogen limitation, more carbon flux is directed to curdlan synthesis with low specific growth rate. When ntrB and ntrC genes in Agrobacterium sp. were inactivated, NH₄Cl utilization ability was significantly impaired in the ntrB and ntrC mutants and curdlan production was significantly reduced. Through proteomic analysis, nearly 40 proteins did not express in ntrC mutant compared with wild type strain. The levels of 22 proteins were significantly increased and 21 proteins were repressed after nitrogen exhaustion. Phosphoglucomutase activity in Agrobacterium sp. was also decreased. However, phosphoglucomutase activity in the ntrC mutant did not change. On that basis, an NtrC-dependent regulatory network for curdlan biosynthesis in response to nitrogen limitation in Agrobacterium sp. ATCC 31749 is proposed.     

马铃薯遗传转化体系的优化及玉米淀粉分支酶基因SBEⅡb的导入

以马铃薯脱毒试管苗茎段为转化受体材料,建立并优化了农杆菌介导的马铃薯遗传转化体系。通过农杆菌介导法将玉米淀粉分支酶基因(Starch branching enzyme b,SBEⅡb)的过表达载体转化马铃薯,接种762个茎段,共获得35株抗性植株。经PCR检测获得了4株转基因阳性植株;对转基因植株进一步进行GUS活性组织化学染色,发现转基因植株的茎段与试管薯均被染上蓝色,表明外源SBEⅡb基因已整合到马铃薯基因组,且正常表达。     

董棕和短穗鱼尾葵光合效率日变化及其与生理生态因子的关系

利用美国产CI-340便携式光合测定系统,对董棕和短穗鱼尾葵的光合日变化特性进行了测定,通过相关分析和通径分析探讨净光合速率与生理生态因子的关系,并利用二项式数学模型对其光合光响应曲线进行了拟合.结果表明:(1)2种棕榈植物的净光合速率日变化都呈现不对称的双峰曲线,有明显的"午休"现象,董棕的午休深度及时长明显大于短穗鱼尾葵,其第二峰出现的时间比短穗鱼尾葵延后2h,这可能主要由于两者在气孔导度及蒸腾速率等自身生理因子活性方面的差异所致.(2)影响董棕和短穗鱼尾葵净光合速率日变化的主要决定生理因子均为胞间CO2浓度,但主要限制因子董棕为气孔导度,短穗鱼尾葵为叶面温度;主要决定生态因子董棕为光合有效辐射,短穗鱼尾葵为空气CO2浓度,主要限定因子两者均为空气相对湿度.(3)董棕和短穗鱼尾葵均具有较高的光补偿点和光饱和点,属强耐阳性植物,但鱼尾葵相对较低,更适于北移引种栽培.     

Molecular phylogeography of Fagus engleriana (Fagaceae) in subtropical China: limited admixture among multiple refugia

Beech is one of the most important trees in the temperate and subtropical forests of the Northern Hemisphere. Despite Chinese beeches have the particularity that only grow in subtropical areas, they have received few phylogeographic research. In this study, we sampled 25 populations of the northernmost-distributed Chinese beech, Fagus engleriana, and detected six haplotypes across 350 individuals by using sequences of two chloroplast intergenic spacers. The chloroplast genetic diversity was relatively low (h _T?=?0.659), with most genetic variance residing among populations (G _ST?=?0.831, N _ST?=?0.855, G _ST??N _ST). SAMOVA analysis indicated that populations clustered into six groups with little admixture among them (most groups were characterized by a unique hapotype). Pairwise difference among haplotypes and Fu??s Fs statistic indicated that populations of F. engleriana have not experienced recent sudden expansions. Both the phylogeographic and demographic patterns found in this study suggest that F. engleriana remained fragmented in multiple refugia throughout the Pleistocene climatic changes, and experienced limited both glacial and interglacial/postglacial expansion. The results of this study imply that long-term isolation among multiple refugia, coupled with little admixture among populations of different refugia provided numerous opportunities for population divergence and allopatric speciation, which might be a driving factor for the exceptionally broad temperate species diversity in southern China.