A computational platform for the high-throughput identification of short RNAs and their targets in plants

细胞中的大多数RNA分子参与蛋白质的生产(核糖体、转运体或信使RNA)。然而，还有一些具有其他功能的RNA分子。最近发现的这类非编码RNA有两类，即microRNAs(MiRNAs)和小干扰RNAs(SiRNAs)。这些调节RNA分子非常短(19-24个核苷酸)，因此通常被称为短RNA(SRNAs)。一些sRNA可以与特定的mRNA相互作用，因为它们具有部分互补的序列。作为这些相互作用的结果，靶向mRNAs的表达显著减少。其他sRNA可以针对染色体并触发DNA修饰。已在植物(拟南芥、水稻和杨树)中鉴定出100多个miRNAs。然而，植物中的sRNAs总数要高得多：最近的一项实验研究发现，在拟南芥中约有75000个sRNAs。此外，在一个物种中发现的几个miRNAs在另一个物种中缺失，这表明存在特定植物群体所特有的miRNAs。为了系统地鉴定植物中的sRNA，联合PIS实验室已经开始使用一种新的高通量测序技术(454焦磷酸测序)。最初，他们正在研究拟南芥，后来，随着基因组序列的公布，将对番茄和紫花苜蓿的sRNA进行分析。这项新技术为每个样本产生了大约200,000个sRNA序列。454技术的初步结果目前是使用标准生物信息学工具手动处理的。然而，对于将从未来454个实验中获得的数百万个sRNA序列来说，这样的分析是不可行的。该项目的主要目标是开发一个专门用于分析高通量454个SRNA测序项目产生的数据的计算平台。该平台将对新的sRNA进行分类，其中一些将进行进一步的实验工作，并寻找可能的RNA靶标。它将首先在来自拟南芥的454个数据上进行测试，然后在番茄和紫花苜蓿上进行测试。在稍后的项目中，将采用比较分析工具进行突变分析。通过该项目发现的新的生物信息学工具和新颖的sRNA将公之于众。确定不同植物物种中sRNA的完整序列将使我们能够描述特定植物性状中一个重要且鲜为人知的调节层，如番茄的肉质果实发育和成熟，以及紫花苜蓿的固氮。

Most of the RNA molecules in cells are involved in protein production (ribosomal, transfer or messenger RNAs). However there are RNA molecules with other functions. Two classes of such non-coding RNAs which have been recently discovered are microRNAs (miRNAs) and small interfering RNAs (siRNAs). These regulatory RNA molecules are very short (19-24 nucleotides) and are thus commonly known as short RNAs (sRNAs). Some sRNAs can interact with specific mRNAs because they have partially complementary sequences. As a result of these interactions the expression of the targeted mRNAs is significantly reduced. Other sRNAs can target the chromosomes and trigger DNA modification. More than a hundred miRNAs have been identified in plant species (Arabidopsis, rice and poplar). However, the total number of sRNAs in plants is much higher: A recent experimental study identified about 75,000 sRNAs in Arabidopsis. In addition, several miRNAs found in one species were absent from the other suggesting that there are miRNAs which are specific to certain groups of plants. To systematically identify sRNAs in plants, the co-PIs laboratories have started to use a novel high-throughput sequencing technology (454 pyrosequencing). Initially, they are studying Arabidopsis and later, as the genome sequences will become available, tomato and alfalfa sRNAs will be analysed. This novel technology produces about 200,000 sRNA sequences for each sample. Preliminary results from the 454 technology are currently processed manually using standard bioinformatics tools. However, such an analysis is unfeasible for the millions of sRNA sequences that will be derived from future 454 experiments. The main goal of this project is to develop a computational platform dedicated to the analysis of data generated by the high-throughput 454 sRNA sequencing projects. This platform will classify new sRNAs, some of which will be subjected to further experimental work, and search for possible RNA targets. It will be initially tested on 454 data from Arabidopis, and subsequently on tomato and alfalfa. Later in the project, a comparative analysis tools will be incorporated for mutant analysis. New bioinformatics tools and novel sRNAs discovered through this project will be made publically available. Identifying the full complement of sRNAs in different plant species will allow us to characterise an important and little understood layer of regulation in specific plant traits such as fleshy fruit development and ripening in tomato, and nitrogen fixation in alfalfa.