An Ac transposon system based on maize chromosome 4S for isolating long-distance-transposed Ac tags in the maize genome

Transposon tagging is an important tool for gene isolation and functional studies. In maize, several transposon-tagging systems have been developed, mostly using Activator/Dissociation (Ac/Ds) and Mutator systems. Here, we establish another Ac-based transposon system with the donor Ac tightly linked with sugary1 (su1) on maize chromosome 4S. Newly transposed Ac (tr-Acs) were detected based on a negative dosage effect, and long-distance-transposed Ac events were identified and isolated from the donor Ac by a simple backcross scheme. In this study, we identified 208 independent long-distance-transposed Ac lines. Thirty-one flanking sequences of these tr-Acs were isolated and localized in the maize genome. As found in previous studies, the tr-Acs preferentially inserted into genic sequences. The distribution of tr-Acs is not random. In our study, the tr-Acs preferentially transposed into chromosomes 1, 2, 9 and 10. We discuss the preferential distribution of tr-Acs from Ac systems. Our system is complementary to two other Ac-based regional-mutagenesis systems in maize, and the combined use of these systems will achieve an even and high-density distribution of Ac elements throughout the maize genome for functional-genomics studies.     

A novel deep learning method for maize disease identification based on small sample-size and complex background datasets

Maize diseases are a major source of yield loss, but due to the lack of human experience and limitations of traditional image-recognition technology, obtaining satisfactory large-scale identification results of maize diseases are difficult. Fortunately, the advancement of deep learning-based technology makes it possible to automatically identify diseases. However, it still faces issues caused by small sample sizes and complex field background, which affect the accuracy of disease identification. To address these issues, a deep learning-based method was proposed for maize disease identification in this paper. DenseNet121 was used as the main extraction network and a multi-dilated-CBAM-DenseNet (MDCDenseNet) model was built by combining the multi-dilated module and convolutional block attention module (CBAM) attention mechanism. Five models of MDCDenseNet, DenseNet121, ResNet50, MobileNetV2, and NASNetMobile were compared and tested using three kinds of maize leave images from the PlantVillage dataset and field-collected at Northeast Agricultural University in China. Furthermore, auxiliary classifier generative adversarial network (ACGAN) and transfer learning were used to expand the dataset and pre-train for optimal identification results. When tested on field-collected datasets with a complex background, the MDCDenseNet model outperformed compared to these models with an accuracy of 98.84%. Therefore, it can provide a viable reference for the identification of maize leaf diseases collected from the farmland with a small sample size and complex background.     

Gene coexpression network alignment and conservation of gene modules between two grass species: maize and rice

One major objective for plant biology is the discovery of molecular subsystems underlying complex traits. The use of genetic and genomic resources combined in a systems genetics approach offers a means for approaching this goal. This study describes a maize (Zea mays) gene coexpression network built from publicly available expression arrays. The maize network consisted of 2,071 loci that were divided into 34 distinct modules that contained 1,928 enriched functional annotation terms and 35 cofunctional gene clusters. Of note, 391 maize genes of unknown function were found to be coexpressed within modules along with genes of known function. A global network alignment was made between this maize network and a previously described rice (Oryza sativa) coexpression network. The IsoRankN tool was used, which incorporates both gene homology and network topology for the alignment. A total of 1,173 aligned loci were detected between the two grass networks, which condensed into 154 conserved subgraphs that preserved 4,758 coexpression edges in rice and 6,105 coexpression edges in maize. This study provides an early view into maize coexpression space and provides an initial network-based framework for the translation of functional genomic and genetic information between these two vital agricultural species.     

Is My Network Module Preserved and Reproducible?

In many applications, one is interested in determining which of the properties of a network module change across conditions. For example, to validate the existence of a module, it is desirable to show that it is reproducible (or preserved) in an independent test network. Here we study several types of network preservation statistics that do not require a module assignment in the test network. We distinguish network preservation statistics by the type of the underlying network. Some preservation statistics are defined for a general network (defined by an adjacency matrix) while others are only defined for a correlation network (constructed on the basis of pairwise correlations between numeric variables). Our applications show that the correlation structure facilitates the definition of particularly powerful module preservation statistics. We illustrate that evaluating module preservation is in general different from evaluating cluster preservation. We find that it is advantageous to aggregate multiple preservation statistics into summary preservation statistics. We illustrate the use of these methods in six gene co-expression network applications including 1) preservation of cholesterol biosynthesis pathway in mouse tissues, 2) comparison of human and chimpanzee brain networks, 3) preservation of selected KEGG pathways between human and chimpanzee brain networks, 4) sex differences in human cortical networks, 5) sex differences in mouse liver networks. While we find no evidence for sex specific modules in human cortical networks, we find that several human cortical modules are less preserved in chimpanzees. In particular, apoptosis genes are differentially co-expressed between humans and chimpanzees. Our simulation studies and applications show that module preservation statistics are useful for studying differences between the modular structure of networks. Data, R software and accompanying tutorials can be downloaded from the following webpage: http://www.genetics.ucla.edu/labs/horvath/CoexpressionNetwork/ModulePreservation.     

A predicted protein-protein interaction network of the filamentous fungus Neurospora crassa

The filamentous fungus Neurospora crassa is a leading model organism for circadian clock studies. Computational identification of a protein-protein interaction (PPI) network (also known as an interactome) in N. crassa can provide new insights into the cellular functions of proteins. Using two well-established bioinformatics methods (the interolog method and the domain interaction-based method), we predicted 27,588 PPIs among 3006 N. crassa proteins. To the best of our knowledge, this is the first identified interactome for N. crassa, although it remains problematic because of incomplete interactions and false positives. In particular, the established PPI network has provided clues to further decipher the molecular mechanism of circadian rhythmicity. For instance, we found that clock-controlled genes (ccgs) are more likely to act as bottlenecks in the established PPI network. We also identified an important module related to circadian oscillators, and some functional unknown proteins in this module may serve as potential candidates for new oscillators. Finally, all predicted PPIs were compiled into a user-friendly database server (NCPI), which is freely available at .     

RNA Silencing Mediated by Direct Repeats in Maize: A Potential Tool for Functional Genomics

It has been shown in tobacco and Arabidopsis that transgenes with multiple direct repeats induce RNA silencing at high frequency. In this study, we tried to establish a direct repeat-induced RNA silencing system in maize and evaluate whether it can be developed as a high throughput tool for functional genomics. Our results showed that the construct phC4, which carries four direct repeats of a chloramphenicol acetyl-transferase (CAT) gene, was able to induce silencing of itself with high efficiency in maize. Using a transient expression system, we further demonstrated that construct phC3G with a β-glucuronidase (GUS) gene located downstream of three direct repeats of CAT gene silenced not only itself in maize calli but also an “endogenous” GUS gene, which was stably expressed in maize calli. Most importantly, when constructs with the maize iojap (ij) gene inserted in either sense or antisense orientation into the downstream of four direct repeats of CAT gene were transformed into maize plants, co-suppression of endogenous and transgenic ij genes was detected in majority of transgenic maize plants. Our co-suppression results suggest that with improvements, this new approach has the potential to become an efficient research tool for high throughput functional genomics.     

基于网络的肾癌随访系统的建立与应用

目的:肾癌是泌尿系常见肿瘤,需长期随访,传统统计及随访方式已逐渐不能适应现代临床及科研需求,我们尝试建立基于网络的肾癌随访系统,满足临床实际工作和科研需求。方法:以Microsoft Office Access 2007软件构建数据库管理系统,分为资料录入模块、检索分析模块、网络随访模块,选取我院2008.5至2012.12收治的132例肾癌患者,将资料录入已建成的数据库,利用网络随访辅助传统随访方式进行随访。结果:构建的肾癌随访数据库,能够实现授权账户登录,检索、分析、肿瘤自动评级等功能,可根据不同检索条件筛选出适合病历,在传统随访方式的基础上联合应用网络随访方式可提高随访效率及随访率,并能获得更详实的随访资料。结论:基于网络的肾癌随访系统操作简便,能够实现无纸化管理,数据详实,检索便捷,为科学制定治疗方案提供帮助,可一定程度上提高随访率及随访效率,降低失访率和患者就医成本。     

玉米转基因技术专利权人合作态势分析

转基因玉米是最重要的转基因主粮作物之一,其转基因技术具有一定的代表性。为了更好地了解和掌握玉米转基因技术领域研发主体合作情况,文章构建了基于专利权人合作网络的目标技术领域专利权人合作态势分析框架,并基于社会网络分析方法与技术,以世界范围内的转基因玉米领域的重要专利权人为分析对象,构建合作网络、分析整体合作特征、挖掘合作子网、探测领域内重要专利权人,从而从宏观、中观和微观三个层面客观展现玉米转基因技术领域专利权人合作态势,为科技战略规划提供一定的决策支撑。     

基因调控网络的模块化组织研究

基因调控网络表现的是大量基因受到转录因子的调控而最终转录翻译为蛋白质进而实现生物功能的复杂信息, 是人们理解生物过程和基因功能的重要内容。为了理解基因调控网络中的调控机理, 网络的拓扑结构及其组织方式是极其重要的研究内容之一。它不仅能说明网络的局部特征, 并且能揭示调控网络的构造方法, 同时还能对调控信号通路进行全面系统的分析。调控网络可分为4层结构: 调控元件、Motif、模块和整个网络。当前, 这种层次结构受到人们越来越多的认可。文中重点讨论motif和模块两层, 比较分析了近年来对网络组织结构的多方面研究内容, 阐述了各个研究结果与结论具有的生物学意义, 并指出了其中存在的问题。在此基础上, 文中还针对这些问题提出了可能存在的研究方向, 并展望了基因调控网络模块化组织的研究前景。     

基于生命周期视角的产业资源生态管理效益分析——以虚拟共生网络系统为例

资源流代谢失调是造成产业生态环境问题的主要原因之一,对其实施基于共生网络的生态管理是解决问题的一项重要的举措。运用全生命周期的思想构建了产业资源共生网络及其管理框架,并运用全生命周期评价的方法,借助生命周期评价软件GaBi4,分别选取EI99 (Eco-Indicator 99)、CML2001 EP评价体系,以武汉市造纸产业为例,通过设计合理的资源流网络关系及中水、废纸和污泥利用共生路径构建虚拟造纸产业共生网络,对比分析了共生设计系统与原有共生系统的各生态环境影响。并运用市场价值法对共生设计系统的经济效益进行了分析。结果表明:共生设计系统总的环境影响、生态系统质量、人体健康、资源损耗值的环境影响分值分别为1166.445、814.509、148.893、203.045,比原有系统分别减少23.91%、19.15%、46.56%、22.26%;其中富营养化、气候变化的影响分别比原有系统降低56.25%、16.62%。同时共生设计系统通过污水、废纸及污泥的回用,在不考虑市场波动的情况下,可获得1018-7252万元的经济效益。可见,通过构建共生网络的生态管理是提高资源利用效率的有效手段之一,在一定条件下可取得明显的环境和经济效益。     

Molecular evidence on maize specific DNA fragment transferred into wheat through sexual hybridization

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淀粉分支酶sbeIIb基因RNA干涉片段转化玉米自交系的研究

玉米高直链淀粉育种是玉米分子育种的一个重要研究方向.本实验中,首先研究了不同诱导愈伤培养基对再生体系的影响,确定了LS+2,4-D 2.0 mg/L+L-pro 700 mg/L+CH 500 mg/L+3 %蔗糖为诱导培养基.同时,构建并验证了含有淀粉分支酶sbeIIb基因双干涉片段载体和胚乳特异性启动子的表达载体pCAMBIA 1301+Glu+1620,并转入根癌农杆菌EHA105,以农杆菌转化法转化玉米自交系178.通过PCR检测,5株转化株表现阳性,初步证明了干涉片段已整合入玉米基因组中.     

Stable integration of an engineered megabase repeat array into the maize genome

Plant genome engineering as a practical matter will require stable introduction of long and complex segments of DNA sequence into plant genomes. Here we show that it is possible to synthetically engineer and introduce centromere-sized satellite repeat arrays into maize. We designed a synthetic repeat monomer of 156 bp that contains five DNA-binding motifs (LacO, TetO, Gal4, LexA, and CENPB), and extended it into tandem arrays using an overlapping PCR method similar to that commonly used in gene synthesis. The PCR products were then directly transformed into maize using biolistic transformation. We identified three resulting insertion sites (arrayed binding sites), the longest of which is at least 1100 kb. The LacI DNA-binding module is sufficient to efficiently tether YFP to the arrayed binding sites. We conclude that synthetic repeats can be delivered into plant cells by omitting passage through Escherichia coli, that they generally insert into one locus, and that great lengths may be achieved. It is anticipated that these experimental approaches will be useful for future applications in artificial chromosome design.     

Efficient Targeted Genome Modification in Maize Using CRISPR/Cas9 System

CRISPR(clustered regularly interspaced short palindromic repeats)/Cas9 system, which is a newly developed technology for targeted genome modification, has been successfully used in a number of species. In this study, we applied this technology to carry out targeted genome modification in maize. A marker gene Zmzb7 was chosen for targeting. The sg RNA-Cas9 construct was transformed into maize protoplasts, and indel(insertion and deletion) mutations could be detected. A mutant seedling with an expected albino phenotype was obtained from screening 120 seedlings generated from 10 callus events. Mutation efficiency in maize heterochromatic regions was also investigated. Twelve sites with different expression levels in maize centromeres or pericentromere regions were selected. The sg RNACas9 constructs were transformed into protoplasts followed by sequencing the transformed protoplast genomic DNA. The results show that the genes in heterochromatic regions could be targeted by the CRISPR/Cas9 system efficiently, no matter whether they are expressed or not. Meanwhile, off-target mutations were not found in the similar sites having no PAM(protospacer adjacent motif) or having more than two mismatches. Together, our results show that the CRISPR/Cas9 system is a robust and efficient tool for genome modification in both euchromatic and heterochromatic regions in maize.