Generation of the iBeetle templates, development of the iBeetle-base to a central gene function database and mining the data for new relationships by clustering

作为全基因组RNAi筛选的先决条件，将建立所谓的iBeetle-Library，旨在分别在一个拷贝中代表Tribolium基因组的所有基因。我们希望基于EST方法的第一轮模板生产，其中随机克隆的cdna被测序。通过生物信息学序列分析，每个聚类选择一个序列，并将相应的质粒整合到iBeetle-Library中。使用基于EST的方法的原因是我们希望尽可能地使用已确认的转录本。此外，我们不希望选择具有异常结构的基因，这些基因可能在自动注释中被遗漏。最后，当前版本的基因组序列还没有达到果蝇或秀丽隐杆线虫对应的质量。因此，一种完全基于PCR的方法——就像在这些物种中所做的那样——会错过重要的基因，会导致重复注射同一基因（因为许多基因被错误地预测为两个），而且当引物位于错误预测的外显子时，容易出现技术问题。为了包括ESTs未涵盖的低表达水平基因，我们将通过基于PCR的方法进一步扩展iBeetle-Library。作为先决条件，我们将通过最先进的测序方法产生转录组数据来增强基因组序列的注释（与社区中的其他人合作）。基于iBeetle-Library，双链rna被用于基因的全基因组敲除。此外，这将是第一次全面收集Tribolium基因，这可能是进一步项目的基础，基因组注释的增强将加强对Tribolium的研究。此外，还将生成一个数据库，用于以下几个目的：1)筛选：在筛选过程中，数据将通过web界面收集并存储在位于al Göttingen的中央数据库中。数据将以按钮、带有预定义词汇表的下拉菜单、自由文本字段和图片的形式收集。此外，还可以通过输入的数据来监控屏幕的进度和效率。2)全基因组表型信息分析。3)发表和与其他资源的链接：数据将与其他数据库（BeetleBase， FlyBase等）链接，广泛的在线搜索工具将允许为联盟识别表型，并在发表后为整个社区识别表型。

As prerequisite for the genome wide RNAi screen, the so called iBeetle-Library will be established that aims at representing all genes of the Tribolium genome in one copy, respectively. We want to base the first round of template production on an EST approach, where randomly cloned cDNAs are sequenced. By bioinformatics sequence analysis, one sequence per cluster will be selected and the respective plasmid integrated into the iBeetle-Library. The reason for the EST based approach is that we want to work with confirmed transcripts as far as possible. Moreover, we do not want to select against genes with unusual structure that might be missed in automatic annotations. Finally, the current version of the genomic sequence has not yet reached the quality of the Drosophila or C.elegans counterparts. Therefore, an entirely PCR based approach - as was performed in these species - would miss important genes, would lead to duplicate injections of the same gene (because many genes are erroneously predicted as two) and would be prone to technical problems when primers are located in erroneously predicted exons. In order to also include genes with low expression level that are not covered by the ESTs, we will further expand the iBeetle-Library by a PCR based approach. As prerequisite for this, we will enhance the annotation of the genomic sequence (in collaboration with others in the community) by producing transcriptome data by state of the art sequencing methods. Based on the iBeetle-Library, double stranded RNAs are produced for the genome wide knock-down of genes. Furthermore, this wili bc the first comprehensive collection of Tribolium genes that is likely to be the basis for further projects and the enhancement of the genome annotation will enhance the research on Tribolium beyond this project.In addition, a database will be generated that serves several aims: I) Screening: During the screen, the data will be collected via web interface and stored in the central database located al Göttingen. The data will be collected in form of buttons, drop-down menus with a predefined vocabulary, free text fields and pictures. Moreover, the progress and efficiency of the screen can be monitored by using the entered data. 2) Analysis of genome wide phenotypic information. 3) Publication and linking with other resources: The data will be linked lo other databases (BeetleBase, FlyBase etc) and an extensive online search tool will allow identifying phenotypes for the consortium and - upon publication - for the entire community.