白菜型冬油菜HDACs基因家族的鉴定及表达分析北大核心CSCD

为了研究HDACs(组蛋白去乙酰化酶)在白菜型冬油菜响应低温胁迫中的影响,该研究采用强抗寒品种‘陇油7号’、耐寒品种‘天油4号’和弱抗寒品种Lenox等白菜型冬油菜,分析HDACs抑制剂曲古抑菌素A(TSA)处理后白菜型冬油菜种子的萌发,及低温胁迫下抗氧化酶活性和渗透调节物质含量变化,并全基因组鉴定HDACs的基因家族,分析其在不同抗寒性白菜型冬油菜中的表达特性。结果表明:(1)1μmol/L和3μmol/L的TSA对3个品种的根长均具有抑制作用,不同温度下外源喷施TSA均可增加抗氧化酶活性和渗透调节物质含量。(2)在白菜型冬油菜中共鉴定出21个HDACs;系统发育分析显示,HDACs可分为3个亚家族,所有基因不均等分布在8条染色体上,但7号和8号染色体上没有基因分布;HDACs启动子序列中包含胁迫响应、光响应、激素应答等相关的作用元件。(3)组织表达结果显示,大部分基因在生长锥中表达量明显高于其他组织部位,BrapaHDA19、BrapaHDA6-1、BrapaHDA8和BrapaHDA7-2在白菜型冬油菜花中表达量较高,且BrapaHDA5-2在‘天油4号’的茎生叶、角果、新基叶和老基叶中的表达量较高,而BrapaHDA7-2在Lenox的花中表达量较高。(4)低温胁迫表达结果显示,BrapaHDA8、BrapaHDA14、BrapaHDA2、BrapaHDT2-2和BrapaHDA15-1在3个抗寒性不同的材料中呈现出差异表达。研究推测,所鉴定的这些基因可能在白菜型冬油菜生长发育及抗寒性方面发挥重要功能。     

甘草质膜水通道蛋白GuPIP1对甘油的转运

旨在进行甘草质膜水通道蛋白GuPIP1对水和溶质通透性的鉴定.将GuPIP1全长编码区的cDNA构入非洲爪蟾卵母细胞检测系统的表达栽体psp64 Poly(A),并将体外转录获得GuPI1的cRNA显微注射入卵母细胞,使GuPIP1基因在卵母细胞中表达,通过测定卵母细胞对水和溶质的转运功能来鉴定GuPIP1的转运功能.结果表明,GuPIP1在异源表达系统非洲爪蟾卵母细胞中具水和甘油通透性,但不能转运尿素,为探讨GuPIP1在植株生理中的作用奠定基础.     

In vitro and in vivo evaluation of O-alkyl derivatives of tramadol

Tramadol is a centrally acting opioid analgesic structurally related to codeine and morphine. O-Alkyl, N-desmethyl, and non-phenol containing derivatives of tramadol were synthesized to probe their effect on metabolic stability and both in vitro and in vivo potency.     

Splenocytes Seed Bone Marrow of Myeloablated Mice: Implication for Atherosclerosis

Extramedullary hematopoiesis has been shown to contribute to the pathogenesis of a variety of diseases including cardiovascular diseases. In this process, the spleen is seeded with mobilized bone marrow cells that augment its hematopoietic ability. It is unclear whether these immigrant cells that are produced/reprogrammed in spleen are similar or different from those found in the bone marrow. To begin to understand this, we investigated the relative potency of adult splenocytes per se to repopulate bone marrow of lethally-irradiated mice and its functional consequences in atherosclerosis. The splenocytes were harvested from GFP donor mice and transplanted into myeloablated wild type recipient mice without the inclusion of any bone marrow helper cells. We found that adult splenocytes repopulated bone marrow of myeloablated mice and the transplanted cells differentiated into a full repertoire of myeloid cell lineages. The level of monocytes/macrophages in the bone marrow of recipient mice was dependent on the cell origin, i.e., the donor splenocytes gave rise to significantly more monocytes/macrophages than the donor bone marrow cells. This occurred despite a significantly lower number of hematopoietic stem cells being present in the donor splenocytes when compared with donor bone marrow cells. Atherosclerosis studies revealed that donor splenocytes displayed a similar level of atherogenic and atheroprotective activities to those of donor bone marrow cells. Cell culture studies showed that the phenotype of macrophages derived from spleen is different from those of bone marrow. Together, these results demonstrate that splenocytes can seed bone marrow of myeloablated mice and modulate atherosclerosis. In addition, our study shows the potential of splenocytes for therapeutic interventions in inflammatory disease.     

One-step isolation of adenosine triphosphate from crude fermentation broth of Saccharomyces cerevisiae by anion-exchange chromatography using supermacroporous cryogel

Adenosine triphosphate (ATP) is an important high-energy compound widely used in biological and therapeutic fields. It can be produced by phosphorylation of adenosine monophosphate (AMP) with microbial cells in industrial scale and the effective isolation of ATP from microbial fermentation broth is a challenging work. In this work, we develop a novel one-step method to directly separate ATP from fermentation broth of Saccharomyces cerevisiae by anion-exchange chromatography using supermacroporous cryogel. The cryogel bed with tertiary amine groups was prepared by grafting N,N-dimethylaminoethyl methacrylate (DMAEMA) monomer chains onto the matrix of a polyacrylamide-based cryogel in a glass column and its properties of liquid dispersion, water permeability, porosity as well as the ligand density were measured. Chromatographic separation of ATP from the fermentation broth by the cryogel was carried out using deionised water and 0.01 M HCl as running buffer, respectively. The breakthrough characteristics and elution performance in the cryogel bed were revealed and analyzed. The purities of the obtained ATP were analyzed quantitatively by high performance liquid chromatography (HPLC). The maximal purity of ATP by the one-step separation method was 95.5% using 0.01 M HCl as running buffer in this work. The corresponding chromatographic behaviors were investigated and analyzed.     

Improving the prediction of RNA secondary structure by detecting and assessing conserved stems

The prediction of RNA secondary structure can be facilitated by incorporating with comparative analysis of homologous sequences. However, most of existing comparative methods are vulnerable to alignment errors and thus are of low accuracy in practical application. Here we improve the prediction of RNA secondary structure by detecting and assessing conserved stems shared by all sequences in the alignment. Our method can be summarized by: 1) we detect possible stems in single RNA sequence using the so-called position matrix with which some possibly paired positions can be uncovered; 2) we detect conserved stems across multiple RNA sequences by multiplying the position matrices; 3) we assess the conserved stems using the Signal-to-Noise; 4) we compute the optimized secondary structure by incorporating the so-called reliable conserved stems with predictions by RNAalifold program. We tested our method on data sets of RNA alignments with known secondary structures. The accuracy, measured as sensitivity and specificity, of our method is greater than predictions by RNAalifold.     

表达狂犬病毒糖蛋白抗原的重组犬2型腺病毒的构建及鉴定

为研制一种预防犬科动物狂犬病的新型疫苗,将含有狂犬病毒ERA株糖蛋白基因(Rabies glycoprotein,Rgp)表达盒的穿梭质粒pVAXΔE3Rgp中的Rgp表达盒克隆入犬2型腺病毒(Canine adenovirus type2,CAV2)骨架质粒pPoly2-CAV2中,获得重组质粒pPoly2-CAV2-ΔE3-Rgp,释放其基因组,转染MDCK细胞系,获得E3缺失区(Deletion of early protein3,ΔE3)含有Rgp表达盒的重组病毒CAV2-ΔE3-Rgp。该重组病毒能在MDCK细胞上产生典型的腺病毒细胞病变。通过酶切、PCR、基因测序,表明该重组病毒含有完整的Rgp表达盒。通过RT-PCR、Western blot等检测,表明该重组病毒能够表达Rgp抗原。用该重组病毒免疫犬,3次免疫后,可以诱导犬产生特异的抗CAV2HI抗体,其效价超过1∶256和抗狂犬病病毒(Rabies virus,RV)中和抗体,其效价超过0.50IU/mL。试验结果表明,获得的重组病毒免疫犬后,能够产生抗狂犬病毒和腺病毒的高效价保护性抗体,是一种有潜力的犬科动物狂犬病毒腺病毒二联疫苗候选株。     

胎盘型碱性磷酸酶的纯化及部分性质的研究

 <正> 胎盘型碱性磷酸酶(P-ALP)是碱性磷酸酶(EC3.1.3.1.ALP)的一种同工酶。P-ALP除出现于妊娠妇女血清外,一些学者还从恶性肿瘤患者血清中发现此酶,并证明它是一种特异性较强的肿瘤标志物。据此,建立P-ALP的简易纯化方法,制备纯度较高的酶制品是建立P-ALP灵敏的微量检测法的先决条件。本文报道以L-苯丙氨酸(L-phe)为配基,用氯代环氧丙烷活化Sepharose 4B的亲和层析法,从人胎盘纯化P-ALP,并对其部分性质进行了研究。     

Multiple hybridization origin of Ranunculus cantoniensis (4x): evidence from trnL-F and ITS sequences and fluorescent in situ hybridization (FISH)

ITS sequences of Ranunculus cantoniensis apparently an allotetraploid were polymorphic at ten nucleotide sites. ITS-based phylogeny of the complex and its allied species showed that ITS clones of the tetraploid were clustered with R. silerifolius var. silerifolius, R. chinensis, R. silerifolius var. dolicathus and R. trigonus. Chloroplast trnL-F phylogeny showed that the complex is a natural group, in which the tetraploid shared the same clade with R. silerifolius var. dolicathus and R. silerifolius var. silerifolius, whose genetic distances were zero. rDNA FISH showed that the longest rDNA-chromosome of the tetraploid was similar to that of R. silerifolius var. dolicathus exclusively. Combining trnL-F, ITS and FISH data, it is suggested that the most probable parents of the tetraploid were R. silerifolius var. silerifolius, R. chinensis and R. silerifolius var. dolicathus, among them R. silerifolius var. silerifolius donated most. Evidences from DNA sequences and chromosome FISH indicated that the tetraploid was most probably a homoploid hybrid. Thus, a scenario of the tetraploid formation is proposed: the tetraploid was synthesized by two rounds of hybridization. The first round was between two pairs of diploids, forming two tetraploids. The second round was between the two primary tetraploids, producing the allotetraploid, R. cantoniensis, eventually.