GarlicESTdb: an online database and mining tool for garlic EST sequences

Abstract

BackgroundAllium sativum., commonly known as garlic, is a species in the onion genus (Allium), which is a large and diverse one containing over 1,250 species. Its close relatives include chives, onion, leek and shallot. Garlic has been used throughout recorded history for culinary, medicinal use and health benefits. Currently, the interest in garlic is highly increasing due to nutritional and pharmaceutical value including high blood pressure and cholesterol, atherosclerosis and cancer. For all that, there are no comprehensive databases available for Expressed Sequence Tags(EST) of garlic for gene discovery and future efforts of genome annotation. That is why we developed a new garlic database and applications to enable comprehensive analysis of garlic gene expression.DescriptionGarlicESTdb is an integrated database and mining tool for large-scale garlic (Allium sativum) EST sequencing. A total of 21,595 ESTs collected from an in-house cDNA library were used to construct the database. The analysis pipeline is an automated system written in JAVA and consists of the following components: automatic preprocessing of EST reads, assembly of raw sequences, annotation of the assembled sequences, storage of the analyzed information into MySQL databases, and graphic display of all processed data. A web application was implemented with the latest J2EE (Java 2 Platform Enterprise Edition) software technology (JSP/EJB/JavaServlet) for browsing and querying the database, for creation of dynamic web pages on the client side, and for mapping annotated enzymes to KEGG pathways, the AJAX framework was also used partially. The online resources, such as putative annotation, single nucleotide polymorphisms (SNP) and tandem repeat data sets, can be searched by text, explored on the website, searched using BLAST, and downloaded. To archive more significant BLAST results, a curation system was introduced with which biologists can easily edit best-hit annotation information for others to view. The GarlicESTdb web application is freely available at .ConclusionGarlicESTdb is the first incorporated online information database of EST sequences isolated from garlic that can be freely accessed and downloaded. It has many useful features for interactive mining of EST contigs and datasets from each library, including curation of annotated information, expression profiling, information retrieval, and summary of statistics of functional annotation. Consequently, the development of GarlicESTdb will provide a crucial contribution to biologists for data-mining and more efficient experimental studies.

Development and validation of new SSR markers from expressed regions in the garlic genome

“…The DNA concentration of each sample was measured using a Qubit Fluorometer and adjusted to 30 ng µL -1 in tris-ethylene diamine tetraacetic acid (TE) buffer. SSR motifs of 2-6 nucleotide repeats in 7720 nonredundant garlic EST sequences at the publicly available GarlicESTdb (http://garlicdb.kribb.re.kr) (Kim et al, 2009) and in ESTs derived from cDNA-AFLP fragments (unpublished EST sequences ) were screened using SSR Locator v.1 computer software . Primer pairs were designed from the flanking sequences to SSR repeat motifs using the SSR Locater program.…”

Microsatellite marker based analysis of genetic diversity in short day tropical Indian onion and cross amplification in related Allium spp.

“…Fischer and Bachmann (2000) were the first to develop and use genomic microsatellite primers in onion and allied species followed by development of expressed sequence tag (EST) derived microsatellites (Kuhl et al 2004;Martin et al 2005) and their utilisation for mapping (McCallum et al 2006) and cultivar discrimination McCallum et al 2008). Microsatellites have also been identified and characterised in Allium fistulosum L. (Song et al 2004;Tsukazaki et al 2007), garlic (Kim et al 2009) and tested for their applicability in onion and related wild species (Araki et al 2010). Despite so many reports, molecular marker application and diversity assessment in Indian bulb onions is still in its nascent stage.…”

Garlic γ-glutamyl transpeptidases that catalyze deglutamylation of biosynthetic intermediate of alliin

“…Reverse transcription (RT) was performed using SuperScript II reverse transcriptase (Life Technologies) and oligo-d(T) 12-18 . Partial cDNAs of AsGGT1 and AsGGT2 were amplified by PCR using Ex Taq DNA polymerase (Takara, Tokyo, Japan) and oligonucleotide primers designed from the nucleotide sequences of two garlic EST clones, i.e., EPP005LLAA12S004013 and EPP005LLAA12S003688 in GarlicESTdb ( Kim et al, 2009 1 ): AsGGT1-Core-F (5′-ATCGCCACTTCATATGAACC-3′) and AsGGT1-Core-R (5′-GATAATGCTAGATATGGCTC-3′) for AsGGT1 ; AsGGT2-Core-F (5′-CTCCTCCACATTAATGGAAC-3′) and AsGGT2-Core-R (5′-AAGTGGTCCCAAACATTTGTC-3′) for AsGGT2 . For the amplification of a partial region of AsGGT3 cDNA, degenerate primers designed based on the sequences of conserved regions of known GGTs, GGT-degenerate-F (5′-ATHGTNYTNAAYAAYGARATG-3′) and GGT-degenerate-R (5′-CCNCCYTTNCKNGGRTC-3′), were used.…”