Sequences for two cDNAs encoding Arabidopsis thaliana eukaryotic protein synthesis initiation factor 4A.

Two distinct cDNAs encoding protein synthesis initiation factor 4A (eIF-4A) were isolated from an Arabidopsis thaliana cDNA library and sequenced. The deduced amino acid sequences from the two cDNAs were compared to eIF-4A from tobacco, mouse and Saccharomyces cerevisiae. The putative ATP-binding sites and RNA helicase motifs were identified.     

Sequences for two cDNAs encoding Arabidopsis thaliana eukaryotic protein synthesis initiation factor 4A

Two distinct cDNAs encoding protein synthesis initiation factor 4A (eIF-4A) were isolated from an Arabidopsis thaliana cDNA library and sequenced. The deduced amino acid sequences from the two cDNAs were compared to eIF-4A from tobacco, mouse and Saccharomyces cerevisiae. The putative ATP-binding sites and RNA helicase motifs were identified.     

Seventy microsatellite markers from Persea americana Miller (avocado) expressed sequence tags

Expressed sequence tags for Persea americana Mill. were investigated to expand upon the number of informative microsatellite markers available for avocado. Seventy informative loci were discovered using 24 P. americana var. americana Mill. accessions. The number of alleles detected ranged from two to 17 and averaged 7.1 alleles per locus. These primers successfully amplified products in different varieties of P. americana, hybrids and a related species, Persea schiedeana. These primers will be useful for characterizing germplasm, determining genetic relationships of cultivated accessions, and for marker‐assisted development of root rot‐tolerant P. americana var. americana rootstock material.     

Analysis of expressed sequence tags from oil palm (Elaeis guineensis)

This is the first report of a systematic study of genes expressed by means of expressed sequence tag (EST) analysis in oil palm, a species of the Arecales order, a phylogenetically key clade of monocotyledons that is not widely represented in the sequence databases. Five different cDNA libraries were generated from male and female inflorescences, shoot apices and zygotic embryos and unidirectional systematic sequencing was performed. A total of 2411 valid EST sequences were thus obtained. Cluster analysis enabled the identification of 209 groups of related sequences and 1874 singletons. Putative functions were assigned to 1252 of the set of 2083 non-redundant ESTs obtained. The EST database described here is a first step towards gene discovery and cDNA array-based expression analysis in oil palm.     

Analysis of expressed sequence tags (ESTs) from a normalized cDNA library and isolation of EST simple sequence repeats from the invasive cotton mealybug Phenacoccus solenopsis

The cotton mealybug, Phenacoccus solenopsis Tinsley, is a serious and invasive pest. At present, genetic resources for studying P. solenopsis are limited, and this negatively affects genetic research on the organism and, consequently, translational work to improve management of this pest. In the present study, expressed sequence tags (ESTs) were analyzed from a normalized complementary DNA library of P. solenopsis. In addition, EST‐derived microsatellite loci (also known as simple sequence repeats or SSRs) were isolated and characterized. A total of 1107 high‐quality ESTs were acquired from the library. Clustering and assembly analysis resulted in 785 unigenes, which were classified functionally into 23 categories according to the Gene Ontology database. Seven EST‐based SSR markers were developed in this study and are expected to be useful in characterizing how this invasive species was introduced, as well as providing insights into its genetic microevolution.     

Classification and comparative analysis of Curcuma longa L. expressed sequences tags (ESTs) encoding glycine-rich proteins (GRPs)

Glycine-rich proteins (GRPs) are a group of proteins characterized by their high content of glycine residues often occurring in repetitive blocs. The diverse expression pattern and sub cellular localization of various GRPs suggest their implication in different physiological processes. Several GRPs has been isolated and characterized from different monocots and dicots. However, little or no information is available about the structure and function of GRPs in asexually reproducing plants. In this study, in-silico analysis of expressed sequence tag database resulted in the isolation of fifty-one GRPs from Curcuma longa L., an asexually reproducible plant of great medicinal and economic significance. Phylogenetic analysis grouped the GRPs into four distinct classes based on conserved motifs and nature of glycine-rich repeats. Majority of the isolated GRPs exhibited high homology with known GRPs from other plants that are expressed in response to various stresses. The presence of high structural diversity and signal peptide in some GRPs suggest their diverse physiological role and tissue specific localization. The isolated sequences can be used as a framework for cloning, characterization and expressional analysis of GRPs in response to various biotic and abiotic stresses in Curcuma longa as well as other asexually reproducing plants.     

Identification of expressed sequence tags of watermelon (Citrullus lanatus) leaf at the vegetative stage

A cDNA library was constructed using mRNA prepared from leaves of watermelon [Citrullus lanatus (Thunb.) Matsum&Nakai] at the vegetative stage. Randomly selected cDNA clones were sequenced in order to identify potentially informative genes. Database comparisons indicated that out of the 704 watermelon cDNA clones, 399 clones (56.7 %) revealed a high degree of sequence similarity to genes from other organisms. These expressed sequence tag clones were divided into ten categories depending upon gene function. Since this kind of experiment has not previously been carried out in this genome, random nucleotide sequencing of these cDNAs could contribute considerable information concerning the novel genes in this organism. Received: 10 July 1999 / Revision received: 20 December 1999 / Accepted: 11 January 2000     

Research note: Analysis of expressed sequence tags from the chrysophycean alga Ochromonas danica (Heterokontophyta)

A cDNA library was constructed from the chrysophycean alga, Ochromonas danica E. G. Pringsheim. 5′‐end sequencing of about 600 cDNA clones yielded 476 authentic expressed sequence tags (EST) of which 275 showed significant matches (E‐value <10^?4) to sequences in a public database. The annotation of these ESTs was carried out to assess subcellular localization of the putative proteins using several internet‐accessible prediction programs for subcellular localization. These analyses revealed that putative plastid proteins in Ochromonas possess N‐terminal bipartite presequences with a conserved phenylalanine at the N‐terminus of the predicted transit peptide‐like domains, similar to other ‘red‐lineage’ secondary symbiotic organisms. The examination of sequences of 3′‐UTR revealed that, similarly to chlorophyte algae, UGUAA may represent a putative polyadenylation signal in O. danica.     

Eimeria tenella: identification of secretory and surface proteins from expressed sequence tags

To identify new vaccine candidates, Eimeria tenella expressed sequence tags (ESTs) from public databases were analysed for secretory molecules with an especially developed automated in silico strategy termed DNAsignalP. A total of 12,187 ESTs were clustered into 2881 contigs followed by a blastx search, which resulted in a significant number of E. tenella contigs with homologies to entries in public databases. Amino acid sequences of appropriate homologous proteins were analysed for the occurrence of an N-terminal signal sequence using the algorithm signalP. The resulting list of 84 entries comprised 51 contigs whose deduced proteins showed homologies to proteins of apicomplexan parasites. Based on function or localisation, we selected candidate proteins classified as (i) secreted proteins of Apicomplexa parasites, (ii) secreted enzymes, and (iii) transport and signalling proteins. To verify our strategy experimentally, we used a functional complementation system in yeast. For five selected candidate proteins we found that these were indeed secreted. Our approach thus represents an efficient method to identify secretory and surface proteins out of EST databases.     

Comparative analysis of the Arabidopsis and rice expressed sequence tag (EST) sets

Large numbers of expressed sequence tags (ESTs) have now been generated from a variety of model organisms. In plants, substantial collections of ESTs are available for Arabidopsis and rice, in each case representing significant proportions of the estimated total numbers of genes. Large-scale comparisons of Arabidopsis and rice sequences are especially interesting due to the fact that these two species are representatives of the two subclasses of the flowering plants (Dicotyledonae and Monocotyledonae, respectively). Here we present the results of systematic analysis of the Arabidopsis and rice EST sets. Non-redundant sets of sequences from Arabidopsis and rice were first separately derived and then combined so that gene families in common between the two species could be identified. Our results show that 58% of non-singleton ESTs are derived from genes in gene families common to the two species. These gene families constitute the basis of a core set of higher plant genes.     

Analysis of expressed sequence tags (ESTs) from Lentinula edodes

The 1,031 expressed sequence tags (ESTs) from the basidiomycete Lentinula edodes were generated as a pilot experiment to see distribution of genes expressed in L. edodes. Among them, genes for hydrophobin, which are specifically found in filamentous fungi, were the most frequently obtained ESTs (33 times), suggesting that they are highly expressed in L. edodes. In addition to known hydrophobin 1 and 2 types, our analysis revealed the existence of novel types of hydrophobin, which we named hydrophobin 3, 4, and 5. The second and the third most highly obtained ESTs were phosphatidylserine decarboxylase and formate dehydrogenase, which were obtained eight and seven times, respectively. It should be noted that two important genes (argonaute and RNA-dependent RNA polymerase) involved in the RNAi pathway were found, suggesting a future application for gene knock-down by RNA interference. The 53 ESTs were identical with the sequences already reported in L. edodes. The 433 ESTs were found to show significant sequence similarity (E value <1 x 10(-5)) with the proteins reported (or predicted) in other species. In total, 387,952 bp were sequenced and registered in DDBJ/GenBank (accession number BJ998097-BJ999127).     

Mapping of expressed sequence tags from a porcine early embryonic cDNA library

The goal of this study was to identify and map genes expressed during the elongation phase of embryogenesis in swine. Expressed sequence tags were analysed from a previously described porcine cDNA library prepared from elongating swine embryos. Average insert length of randomly selected clones was approximately 600 bp, with a range from < 100 to > 2500 bp. Single-pass, coding strand sequences from 1132 independent clones were compared with the GenBank non-redundant (nr) database via BLASTN analysis to identify potential porcine homologous of known genes. Among these sequences, 781 (69%) showed significant (score > 300) homology to non- mitochondrial sequences previously deposited in GenBank. Sequences matching interleucin 1 beta and thymosin beta 10 were most frequently observed (24 and 18 clones, respectively), in addition to matches with 310 other distinct genes. No significant match in the GenBank nr database was obtained for 303 sequences. Analysis demonstrated that 151 (50%) had open reading frames (ORF) extending at least 50 codons from the first base of the clone insert. Genetic markers were developed and used to map a subset of 17 genes, selected on the basis of function or of the ability to design primers that successfully amplified porcine genomic DNA, to 10 different porcine chromosomes, providing a set of mapped markers corresponding to genes expressed during conceptus elongation.