Abundance, marker development and genetic mapping of microsatellites from unigenes in Brassica napus

Rapeseed (Brassica napus) is the second most important oil crop in the world after soybean. The repertoire of simple sequence repeat (SSR) markers for rapeseed is limited and warrants a search for a larger number of polymorphic SSRs for germplasm characterization and breeding applications. In this study, a total of 5,310 SSR-containing unigenes were identified from a set of 46,038 B. napus unigenes with an average density of one SSR every 5.75?kb. A set of 1,000 expressed sequence tag (EST)-SSR markers with repeat length ??18?bp were developed and tested for their ability to detect polymorphism among a panel of six rapeseed varieties. Of these SSR markers, 776 markers detected clear amplification products, and 511 displayed polymorphisms among the six varieties. Of these polymorphic markers, 195 EST-SSR markers, corresponding to 233 loci, were integrated into an existing B. napus linkage map. These EST-SSRs were randomly distributed on the 19 linkage groups of B. napus. Of the mapped loci, 166 showed significant homology to Arabidopsis genes. Based on the homology, 44 conserved syntenic blocks were identified between B. napus and Arabidopsis genomes. Most of the syntenic blocks were consistent with the duplication and rearrangement events identified previously. In addition, we also identified three previously unreported blocks in B. napus. A subset of 40 SSRs was used to assess genetic diversity in a collection of 192 rapeseed accessions. The polymorphism information content of these markers ranged from 0.0357 to 0.6753 with an average value of 0.3373. These results indicated that the EST-SSR markers developed in this study are useful for genetic mapping, molecular marker-assisted selection and comparative genomics.     

不同冬季覆盖作物对双季稻田土壤有机碳的影响

以冬闲-双季稻种植模式为对照(CK),分析了黑麦草-双季稻(Ry)、紫云英-双季稻(Mv)、马铃薯-双季稻(Po)和油菜-双季稻(Ra)5种不同种植模式下冬季覆盖作物秸秆还田后对0～5、5～10、10～20 cm土壤有机碳、活性有机碳、碳库管理指数和有机碳储量的影响.结果表明:与CK处理相比,黑麦草、紫云英、马铃薯和油菜秸秆还田处理均提高了稻田0～5、5～10、10～20 cm土壤总有机碳和活性有机碳含量,其中Po处理最高.冬季覆盖作物秸秆还田均提高了稻田不同层次土壤的碳库活度、碳库活度指数、碳库指数和土壤碳库管理指数,其大小顺序均表现为Po>Mv>Ry>Ra>CK.不同冬季覆盖作物秸秆还田处理与CK处理相比均有利于提高稻田不同层次土壤有机碳储量,其中以紫云英秸秆还田的效果最好,黑麦草和马铃薯次之;各处理稻田土壤有机碳储量均随土壤深度的增加而递增.     

长期施肥对双季稻田甲烷排放和关键功能微生物的影响

研究不同施肥措施对双季稻田甲烷(CH_4)排放特征的影响及其微生物学机理,对合理利用及评价不同施肥模式对水稻生长的影响具有重要意义。以长期施肥定位试验田为平台,采用静态箱-气相色谱法对施用化肥(MF:mineral fertilizer alone)、秸秆还田配施化肥(RF:rice residues plus mineral fertilizer)、30%有机肥配施70%化肥(LOM:30%organic matter plus 70%mineral fertilizer)、60%有机肥配施40%化肥(HOM:60%organic matter plus 40%mineral fertilizer)和无肥(CK:without fertilizer)条件下双季稻田CH_4排放及其微生物学机理进行了分析。结果表明,早稻和晚稻生长期,不同施肥处理稻田CH_4排放通量均显著高于CK,表现为HOMLOMRFMFCK。各处理间CH_4总排放量差异达显著水平,其大小顺序与排放通量趋势一致,以HOM处理为最高,比CK处理增加105.56%,其次是LOM和RF处理,分别比CK处理增加72.97%和54.17%。关键功能土壤微生物测定结果表明,早稻和晚稻各个主要生育时期,各处理稻田土壤产甲烷古菌的数量变化范围为(3.18—81.07)×10~3cfu/g,土壤甲烷氧化细菌的数量变化范围为(24.82—379.72)×10~3cfu/g。稻田土壤产甲烷古菌和甲烷氧化细菌数量大小顺序为HOMLOMRFMFCK,各施肥处理均显著高于CK;HOM、LOM、RF处理显著高于MF、CK处理。双季稻田CH_4排放与稻田土壤产甲烷古菌、甲烷氧化细菌数量变化关系密切。采用有机无机肥配施促进了双季稻田生态系统CH_4的排放和关键功能微生物的数量。     

人脐血CD34+细胞在NOD/SCID小鼠免疫重建及其抗HBV作用的初步探讨

目的：探讨NOD／SCID（nonobese diabetic／severe combined immunodeficient）小鼠移植人脐带血（human umbilical cord blood,HUCB）CD34＋细胞后免疫重建的特性。建立hu—NOD／SCID人鼠嵌合模型并观察其人源化免疫细胞在小鼠体内的生长分化特性、存活时间及其对HBV感染的清除作用。方法：1．NOD／SCID小鼠于C0603．5Gy照射后24h内尾静脉输注HUCBCD34＋细胞;2．以流式细胞技术鉴定小鼠外周血中CD45＋,CD3＋,CD19＋,CD56＋等人源化细胞的比例;3．NOD／SCID小鼠于移植后第4wk注射HBV感染者血清并以未移植的NOD／SCID小鼠作为对照,注射同等量的患者血清;4．于感染后1、7、10、15天采血,免疫荧光定量PCR方法分别检测其HBV—DNA含量。结果：1．HUCBCD34＋细胞移植后第2wk,在小鼠外周血中检测出的CD3＋CD8＋T细胞、CD3＋CD4＋T细胞、CD19＋B细胞、CD56＋NK细胞的比例分别为18．6％、16．1％、13．1％和27．8％。各细胞比例随小鼠周龄而变化。所有移植小鼠存活时间均达9wk;2．移植后小鼠感染HBV血清后,病毒仅在感染后第一天检出,随后消失;未移植CD34＋细胞的小鼠外周血HBV—DNA一直维持在103水平：结论：1．NOD／SCID小鼠经射线照射后移植HUCBCD34＋细胞,在不加任何刺激因子的情况下小鼠可以长时间存活并重建免疫;2．hu—NOD／SCID人鼠嵌合模型小鼠免疫成功重建后,对HBV感染有快速的清除作用。     

Association of cortactin and fascin-1 expression in gastric adenocarcinoma: correlation with clinicopathological parameters.

Cortactin and fascin-1 are important factors in tumor progression. We tested the hypothesis that cortactin and fascin-1 expression correlates with clinicopathological parameters of gastric adenocarcinoma. Immunohistochemical analysis of cortactin and fascin-1 was done using tissue microarrays of 100 surgical specimens, including 20 well-differentiated, 20 moderately differentiated, and 60 poorly differentiated gastric adenocarcinomas. Among the 20 well-differentiated gastric adenocarcinomas, 15 cases (75%) showed negative or weak staining (1+); 5 cases (25%) had moderate (2+) or strong (3+) cortactin expression. Among the 60 poorly differentiated gastric adenocarcinomas, more than three-quarters of the cases (76.7%) had moderate or strong cortactin expression; 14 cases (23.3%) had weak staining. Of 20 well-differentiated gastric adenocarcinoma cases, 14 (70%) showed negative or weak staining of fascin-1, whereas nearly one-third (30%) had moderate or strong expression. Among the 60 poorly differentiated gastric adenocarcinomas, 32 (53.3%) exhibited moderate or strong fascin-1 expression; fewer than half of the cases showed negative or weak staining. Higher intensity of cortactin and fascin-1 staining correlated directly with more-advanced cancer stages (TNM) and inversely with survival rates. Our findings suggest the possibility that pharmacological inhibitors of cortactin and fascin-1 activity may slow down tumor progression and prolong survival time in patients with gastric adenocarcinomas.     

Expression and purification of an FGF9 fusion protein in E. coli,and the effects of the FGF9 subfamily on human hepatocellular carcinoma cell proliferation and migration

Fibroblast growth factor (FGF) 9 has oncogenic activity and plays an important role in the development of ovarian, lung, prostate, and gastric cancers. In the present study, with the aim of reducing the cost of utilizing growth factors in cancer research, a simple and efficient method for the preparation of recombinant human (rh)FGF9 in Escherichia coli was established. The rhFGF9 fusion protein (6 × His-TEV-rhFGF9) and the native protein released by tobacco etch virus (TEV) protease were obtained using a Ni-NTA system, with > 95% purity. Both purified forms of rhFGF9, with and without fusion tags, significantly stimulated the proliferation of NIH3T3 cells. The FGF9 subfamily, including FGF9, FGF16, and FGF20, in addition to rhFGF16, rhFGF9, and rhFGF20, were shown to stimulate the proliferation and migration of HuH7 human hepatocellular carcinoma (HCC) cells. Mechanistic studies revealed that the stimulation of HuH7 cell proliferation and migration with rhFGF9 and rhFGF20 were associated with the activation of the extracellular signal-regulated kinase (ERK) and nuclear factor κB (NF-κB) pathways and matrix metalloproteinase-26 (MMP26). Inhibition of the ERK and NF-κB pathways blocked cell migration, and NF-κB was demonstrated to be regulated by ERK. Therefore, the present study demonstrates a simple method for the preparation of biologically active rhFGF9 protein. Furthermore, the results indicate that exogenous rhFGF9- and rhFGF20-activated ERK/NF-κB signal transduction pathways play important roles in the regulation of HCC cell proliferation and migration, and this discovery helps to find the potential for new solutions of the treatment of liver cancer.

     

Plk1 Self-Organization and Priming Phosphorylation of HsCYK-4 at the Spindle Midzone Regulate the Onset of Division in Human Cells

Animal cells initiate cytokinesis in parallel with anaphase onset, when an actomyosin ring assembles and constricts through localized activation of the small GTPase RhoA, giving rise to a cleavage furrow. Furrow formation relies on positional cues provided by anaphase spindle microtubules (MTs), but how such cues are generated remains unclear. Using chemical genetics to achieve both temporal and spatial control, we show that the self-organized delivery of Polo-like kinase 1 (Plk1) to the midzone and its local phosphorylation of a MT-bound substrate are critical for generating this furrow-inducing signal. When Plk1 was active but unable to target itself to this equatorial landmark, both cortical RhoA recruitment and furrow induction failed to occur, thus recapitulating the effects of anaphase-specific Plk1 inhibition. Using tandem mass spectrometry and phosphospecific antibodies, we found that Plk1 binds and directly phosphorylates the HsCYK-4 subunit of centralspindlin (also known as MgcRacGAP) at the midzone. At serine 157, this modification creates a major docking site for the tandem BRCT repeats of the Rho GTP exchange factor Ect2. Cells expressing only a nonphosphorylatable form of HsCYK-4 failed to localize Ect2 at the midzone and were severely impaired in cleavage furrow formation, implying that HsCYK-4 is Plk1's rate-limiting target upstream of RhoA. Conversely, tethering an inhibitor-resistant allele of Plk1 to HsCYK-4 allowed furrows to form despite global inhibition of all other Plk1 molecules in the cell. Our findings illuminate two key mechanisms governing the initiation of cytokinesis in human cells and illustrate the power of chemical genetics to probe such regulation both in time and space.     

麒麟紫葳的组织培养与快速繁殖

1植物名称麒麟紫葳(Radermachera sinica),又名幸福树、菜豆树. 2材料类别植株带芽茎段. 3培养条件芽诱导、分化培养基:(1)MS 6-BA0.5 mg·L-1(单位下同) IBA 0.2;生根培养基:(2)1/2MS IBA 0.5.上述培养基均添加3%蔗糖、0.6%琼脂,pH(5.8 0.1).培养温度为(28 2)℃,光照时间10 h·d-1,光照度为2 000 1x.     

琼胶酶研究进展

琼胶酶是一种多糖水解酶, 根据其降解琼脂糖的作用方式不同, 可以分为a- 琼胶酶(EC 3. 2.1. -) 和b- 琼胶酶(EC 3.2.1.81)。本文结合自己的研究, 从琼胶酶的生物学研究、酶的分类、晶体结构、催化机理以及酶的应用等几个方面综述了琼胶酶的研究进展。     

On foundations of discrete element analysis of contact in diarthrodial joints

Information about the stress distribution on contact surfaces of adjacent bones is indispensable for analysis of arthritis, bone fracture and remodeling. Numerical solution of the contact problem based on the classical approaches of solid mechanics is sophisticated and time-consuming. However, the solution can be essentially simplified on the following physical grounds. The bone contact surfaces are covered with a layer of articular cartilage, which is a soft tissue as compared to the hard bone. The latter allows ignoring the bone compliance in analysis of the contact problem, i.e. rigid bones are considered to interact through a compliant cartilage. Moreover, cartilage shear stresses and strains can be ignored because of the negligible friction between contacting cartilage layers. Thus, the cartilage can be approximated by a set of unilateral compressive springs normal to the bone surface. The forces in the springs can be computed from the equilibrium equations iteratively accounting for the changing contact area. This is the essence of the discrete element analysis (DEA). Despite the success in applications of DEA to various bone contact problems, its classical formulation required experimental validation because the springs approximating the cartilage were assumed linear while the real articular cartilage exhibited non-linear mechanical response in reported tests. Recent experimental results of Ateshian and his co-workers allow for revisiting the classical DEA formulation and establishing the limits of its applicability. In the present work, it is shown that the linear spring model is remarkably valid within a wide range of large deformations of the cartilage. It is also shown how to extend the classical DEA to the case of strong nonlinearity if necessary.