GMATo: A novel tool for the identification and analysis of microsatellites in large genomes

Simple Sequence Repeats (SSR), also called microsatellite, is very useful for genetic marker development and genome application. The increasing whole sequences of more and more large genomes provide sources for SSR mining in silico. However currently existing SSR mining tools can’t process large genomes efficiently and generate no or poor statistics. Genome-wide Microsatellite Analyzing Tool (GMATo) is a novel tool for SSR mining and statistics at genome aspects. It is faster and more accurate than existed tools SSR Locator and MISA. If a DNA sequence was too long, it was chunked to short segments at several Mb followed by motifs generation and searching using Perl powerful pattern match function. Matched loci data from each chunk were then merged to produce final SSR loci information. Only one input file is required which contains raw fasta DNA sequences and output files in tabular format list all SSR loci information and statistical distribution at four classifications. GMATo was programmed in Java and Perl with both graphic and command line interface, either executable alone in platform independent manner with full parameters control. Software GMATo is a powerful tool for complete SSR characterization in genomes at any size.

Availability

The soft GMATo is freely available at http://sourceforge.net/projects/gmato/files/?source=navbar or on contact     

Ser1333 phosphorylation indicates ROCKI activation

Background

Two isoforms of Rho-associated protein kinase (ROCK), ROCKI and ROCKII, play a pivotal role in regulation of cytoskeleton and are involved in multiple cellular processes in mammalian cells. Knockout mice experiments have indicated that the functions of ROCKI and II are probably non-redundant in physiology. However, it is difficult to differentiate the activation status of ROCKI and ROCKII in biological samples. Previously, we have identified phosphorylation site of ROCKII at Ser1366 residue sensitive to ROCK inhibition. We further investigated the activity-dependent phosphorylation site in ROCKI to establish the reagents that can be used to detect their individual activation.

Results

The phosphorylation site of ROCKI sensitive to its inhibition was identified to be the Ser1333 residue. The ROCKI pSer1333-specific antibody does not cross-react with phosphorylated ROCKII. The extent of S1333 phosphorylation of ROCKI correlates with myosin II light chain phosphorylation in cells in response to RhoA stimulation.

Conclusions

Active ROCKI is phosphorylated at Ser1333 site. Antibodies that recognize phospho-Ser1333 of ROCKI and phospho-S1366 residues of ROCKII offer a means to discriminate their individual active status in cells and tissues.     

Sequence-based marker development in wheat: advances and applications to breeding

In the past two decades, the wheat community has made remarkable progress in developing molecular resources for breeding. A wide variety of molecular tools has been established to accelerate genetic and physical mapping for facilitating the efficient identification of molecular markers linked to genes and QTL of agronomic interest. Already, wheat breeders are benefiting from a wide range of techniques to follow the introgression of the most favorable alleles in elite material and develop improved varieties. Breeders soon will be able to take advantage of new technological developments based on Next Generation Sequencing. In this paper, we review the molecular toolbox available to wheat scientists and breeders for performing fundamental genomic studies and breeding. Special emphasis is given on the production and detection of single nucleotide polymorphisms (SNPs) that should enable a step change in saturating the wheat genome for more efficient genetic studies and for the development of new selection methods. The perspectives offered by the access to an ordered full genome sequence for further marker development and enhanced precision breeding is also discussed. Finally, we discuss the advantages and limitations of marker-assisted selection for supporting wheat improvement.     

Sister chromatid exchange in vivo,chromosomal characterization and NORs activity of leukemia cells during 5FU-treatments

Summary A transplantable mouse leukemia model, the leukemia cell of which has a marker chromosome and the XX genome type which differ obviously from their male host cells provides a possibility to precisely identify the leukemia cells among their male host cells cytogenetically. A sister chromatid exchange (SCE) plus chromosomal C-banding technique that we report here is very useful. The SCE frequencies in vivo of both leukemia cells and host cells were twice as high as the normal mouse cells. The higher SCE frequencies of the host cells in the leukemia mice may be due to some toxicities from the leukemia cells or some biological large molecule exchanges between the leukemia cells and the host cells. There was no significant difference in SCE frequencies between cells from the spleen and from the bone marrow of the leukemia mice. The percentages of leukemia cells in both spleen and bone marrow were more than 90% when the mice had been injected with the leukemia cells for five days. The host cells in the leukemia mice did not become leukemia cells. The 5FU-treated leukemia mice survived very well for more than twenty-three days. After the 5FU-treatments, most of the leukemia cells died, subsequently, SCE frequencies decreased to a normal level. Both the number of Ag-NORs per cell and the number of chromosomes bearing Ag-NORs per cell in the leukemia mice decreased to 60% and 40%, respectively, of the level found in normal mouse cells.     

Cluster-based upper body marker models for three-dimensional kinematic analysis: Comparison with an anatomical model and reliability analysis

Quantifying angular joint kinematics of the upper body is a useful method for assessing upper limb function. Joint angles are commonly obtained via motion capture, tracking markers placed on anatomical landmarks. This method is associated with limitations including administrative burden, soft tissue artifacts, and intra- and inter-tester variability. An alternative method involves the tracking of rigid marker clusters affixed to body segments, calibrated relative to anatomical landmarks or known joint angles. The accuracy and reliability of applying this cluster method to the upper body has, however, not been comprehensively explored. Our objective was to compare three different upper body cluster models with an anatomical model, with respect to joint angles and reliability. Non-disabled participants performed two standardized functional upper limb tasks with anatomical and cluster markers applied concurrently. Joint angle curves obtained via the marker clusters with three different calibration methods were compared to those from an anatomical model, and between-session reliability was assessed for all models. The cluster models produced joint angle curves which were comparable to and highly correlated with those from the anatomical model, but exhibited notable offsets and differences in sensitivity for some degrees of freedom. Between-session reliability was comparable between all models, and good for most degrees of freedom. Overall, the cluster models produced reliable joint angles that, however, cannot be used interchangeably with anatomical model outputs to calculate kinematic metrics. Cluster models appear to be an adequate, and possibly advantageous alternative to anatomical models when the objective is to assess trends in movement behavior.     

不同来源大豆同名品种"满仓金"表现型及SSR标记的异同性分析

以国家种质库中保存的不同来源的大豆同名品种满仓金及与其有亲缘关系的种质共28份为实验材料,时其SSR分子标记及农艺性状进行比较分析,目的是检测种质库中保存的同名品种之间的差异程度,为大豆种质资源的保存、研究与利用以及构建大豆棱心种质提供理论依据。研究结果表明,在28份种质23个SSR位点共检测出151个等位变异,利用其中的5个位点可将28份种质区分开。通过对SSR数据及农艺性状的分析比较,发现利用两种不同类型数据对材料进行分析,既有相同的趋势又存在差异,将不同类型数据结合起来使用能比较全面地阐明品种之间的遗传关系。根据结果推测,全国统一编号分别为ZDD00078、ZDD00924的满仓金与黄宝珠和金元有密切的血缘关系,其他不同来源的满仓金之间差异较大,无遗传一致性,也具有保存价值。     

应用RAPD标记检测导入普通小麦中的Elymus rectisetus遗传物质

以72个含16株系的BC2F5（小麦/Elymus rectisetus//小麦）单株为供体材料,用筛选出的12个10碱基随机引物对其进行多态性扩增。以E.rectisetus和Fukuhokomugi(Triticum aestivum)为亲本,建立RAPD标记。实验表明：12个随机引物中,有10个随机引物能够在16个株系的68个单株中分别扩增出普通小麦所没有的E.rectisetus的DNA片段,可分为6个类型,此外,1040-2-5-1和1048-Y3-3-1可能含5种异源染色体。     

培美曲塞联合顺铂化疗对晚期非小细胞肺癌患者疗效及血清肿瘤标志物的影响

目的:探究培美曲塞联合顺铂化疗对晚期非小细胞肺癌患者疗效及血清肿瘤标志物的影响。方法:选取于2012年7月~2016年2月期间我院收治的89例非小细胞肺癌患者为研究对象,采用随机数字法将研究对象分为观察组(45例)和对照组(44例);观察组采用培美曲塞联合顺铂化疗,对照组采用多西他赛联合顺铂化疗,观察并比较两组患者治疗前后细胞角质素片段抗原(CYRAF211)、血清癌胚抗原(CEA)、糖类抗原125(CA125)、神经元特异性烯醇化酶(NSE)表达水平。结果:观察组疗效优于对照组,比较有统计学差异(Z=1.940,P=0.026),观察组治疗的总有效率(55.66%)显著高于对照组(36.37%),差异有统计学意义(χ2=5.432,P=0.034);化疗后,两组CEA、CYFRA21-1、CA125及NSE水平均较化疗前较有显著下降,同时观察组各指标水平均显著低于对照组(P0.05);化疗前,Ⅲ期患者肿瘤标志物CEA、CYFRA21-1、CA125及NSE水平均显著低于Ⅳ期患者(P0.05);化疗后,Ⅲ期和Ⅳ期患者肿瘤标志物CEA、CYFRA21-1、CA125及NSE水平较治疗前均有显著降低(P0.05),且Ⅲ期水平显著低于Ⅳ期(P0.05)。结论:培美曲塞联合顺铂治疗晚期非小细胞肺癌具有显著疗效,血清CEA、CYFRA21-1、CA125及NSE水平经化疗后显著降低,可作为分期和评价化疗疗效的可靠指标。     

Application of multiplex-ready PCR for fluorescence-based SSR genotyping in barley and wheat

Microsatellites (SSRs) are widely used in cereal research, and their use in marker assisted breeding has increased the speed and efficiency of germplasm improvement. Central to the application of SSRs for many purposes are methodologies enabling the low-cost acquisition of large quantities of genetic information for gene and genotype identification. In this study, multiplex-ready PCR was evaluated in barley and bread wheat as an approach for rapid and more automated SSR genotyping on a fluorescence-based DNA fragment analyzer. Multiplex-ready PCR is a method that allows SSR genotyping to be performed using a standardized protocol. The method enables flexible fluorescence labeling of SSRs, generates a relatively constant amount of PCR product for each marker, and has a high amenability to multiplex PCR (the simultaneous amplification of several SSRs in the same reaction). A high (92%) compatibility of published SSRs with multiplex-ready PCR is demonstrated, and the usefulness of the method for large scale genotyping is shown by its application for whole genome marker assisted breeding in barley. A database of more than 2,800 barley and wheat SSRs, and a suite of bio-informatic tools were developed to support the deployment of multiplex-ready PCR for various genetic applications, and are accessible at . Multiplex-ready PCR is broadly applicable to cereal genomics research and marker assisted breeding, and should be transferable to similar analyses of any animal or plant species.     

Identification and validation of RAPD and SCAR markers linked to the gene <Emphasis Type="Italic">Er3</Emphasis> conferring resistance to <Emphasis Type="Italic">Erysiphe pisi</Emphasis> DC in pea

Three genes, er1, er2 and Er3, conferring resistance to powdery mildew (Erysiphe pisi) in pea have been described so far. Because single gene-controlled resistance tends to be overcome by evolution of pathogen virulence, accumulation of several resistance genes into a single cultivar should enhance the durability of the resistance. Molecular markers linked to genes controlling resistance to E. pisi may facilitate gene pyramiding in pea breeding programs. Molecular markers linked to er1 and er2 are available. In the present study, molecular markers linked to Er3 have been obtained. A segregating F₂ population derived from the cross between a breeding line carrying the Er3 gene, and the susceptible cultivar ‘Messire’ was developed and genotyped. Bulk Segregant Analysis (BSA) was used to identify Random Amplified Polymorphic DNA (RAPD) markers linked to Er3. Four RAPD markers linked in coupling phase (OPW04_637, OPC04_640, OPF14_1103, and OPAH06_539) and two in repulsion phase (OPAB01_874 and OPAG05_1240), were identified. Two of these, flanking Er3, were converted to Sequence Characterized Amplified Region (SCAR) markers. The SCAR marker SCW4₆₃₇ co-segregated with the resistant gene, allowing the detection of all the resistant individuals. The SCAR marker SCAB1₈₇₄, in repulsion phase with Er3, was located at 2.8 cM from the gene and, in combination with SCW4₆₃₇, was capable to distinguish homozygous resistant individuals from heterozygous with a high efficiency. In addition, the validation for polymorphism in different genetic backgrounds and advanced breeding material confirmed the utility of both markers in marker-assisted selection.