The genus Acanthamoeba can cause severe infections such as granulomatous amebic encephalitis and amebic keratitis in humans. However, little genomic information of Acanthamoeba has been reported. Here, we constructed Acanthamoeba expressed sequence tags (EST) database (Acanthamoeba EST DB) derived from our 4 kinds of Acanthamoeba cDNA library. The Acanthamoeba EST DB contains 3,897 EST generated from amebae under various conditions of long term in vitro culture, mouse brain passage, or encystation, and downloaded data of Acanthamoeba from National Center for Biotechnology Information (NCBI) and Taxonomically Broad EST Database (TBestDB). The almost reported cDNA/genomic sequences of Acanthamoeba provide stand alone BLAST system with nucleotide (BLAST NT) and amino acid (BLAST AA) sequence database. In BLAST results, each gene links for the significant information including sequence data, gene orthology annotations, relevant references, and a BlastX result. This is the first attempt for construction of Acanthamoeba database with genes expressed in diverse conditions. These data were integrated into a database (http://www.amoeba.or.kr).
Acanthamoeba/*genetics ; Animals ; *Databases, Genetic ; *Expressed Sequence Tags

OBJECTIVETo study the distribution frequency and characteristics of nucleotide repeat of 94 923 ESTs for developing microsatellite primers, providing a theoretical basis and technical support for appropriate conservation and application of sweet wormwood (Artemisia annua).

METHODEST-SSR detection was performed using Perl program MISA. Gene Ontology (GO) annotations were formatted for input into the GOSlim program and the output was parsed to count the occurrence of each GO category. Primer 3 software was used to design 18 pairs primers, amplified products were separated on a 6% denaturing polyacrylamide gel using silver staining.

RESULTBy searching with Active Perl, totally 2 110 SSRs were detected, accounting for 8.6%. The frequency of occurrence of dinucleotide and trinucleotide was 28% and 50.4%, respectively. The most common repeat motifs of trinucleotide were ACC/GGT, accounting for 9.8%. Three hundred and twelve SSR-ESTs were annotated using GO terms. The suitable PCR system of 15 pairs primers was established, and revealed microsatellite polymorphism in 36 individuals.

CONCLUSIONThere are a variety of motifs at EST-SSR locus in sweet wormwood, and more effective amplification and polymorphism in 18 pairs detected primers. Therefore, EST resource is an effective and feasible approach to develop SSR markers, and EST-SSRs will a powerful tool for studies of sweet wormwood genetic resources.

Artemisia annua ; genetics ; Expressed Sequence Tags ; Microsatellite Repeats ; Polymorphism, Genetic

To analysis the SSR loci information in the transcriptome of Cordyceps sinensis and develop SSR molecular markers,MISA(MicroSatellite) software was used to analyze the microsatellites information from 16 875 unigene sequences and SSR primer designed by Primer 3. 0. In total,5 899 SSRs were detected in 4 252 unigene with the distribution frequency of 34. 99%,which was represented by 74 repeat motifs and SSR loci occurred per 7 952 bp in length. In the SSRs,the mono-nucleotide was the most abundant repeat motif(42. 5%),followed by tri-nucleotide(34. 48%),C/G and CCG/CGG were the dominant repeat motifs,respectively. The number of repetitions of the six SSR repeat types was concentrated on 5 to 12 times,and the length was mostly less than 24 bp. A total of 12 282 pairs of primers were screened and selected 20 pairs of primers for validity detection randomly,10 pairs of primers amplified the expected specific bands,and primer P1 has significant polymorphism. Moreover,it was found that unigene containing SSR loci is mainly related to genetic and environmental functions after GO and KEGG annotation. In conclusion,these SSR loci in the transcriptome of O. sinensis are high in frequency,rich in primitive types,high in polymorphism,and highly available,which will provides abundant candidate molecular markers for its genetic diversity analysis,resource identification protection,and gene function research.
Cordyceps/genetics* ; Expressed Sequence Tags ; Microsatellite Repeats ; Polymorphism, Genetic ; Transcriptome

In order to make a series of analyses on the gene expression of Eimeria tenella sensitive strain and anti-maduramycin strain at their different developmental stages, we constructed a mixed cDNA library with unsporulated oocyst, sporulated oocyst, sporozoite and merozoite from Eimeria tenella sensitive strain and antimaduramycin strain (induced by its sensitive strain)respectively. After sequencing reactions, the total 2806 high quality expressed sequence tags (ESTs) of 3' ends were derived from the cDNA library. Results of bioinformatics analysis of all EST data showed that EST sequences assembled 1424 tentative unique transcripts (TUTs) and the redundancy was 49.3%, and that about 83.6% TUTs could not be assigned for functional description. Among the remained annotated genes, infection related proteins and development related proteins such as MIC2 protein, BT1 family protein and some ribosomal protein expressed at high abundant level.
Eimeria tenella ; genetics ; Expressed Sequence Tags ; Gene Library ; Sequence Tagged Sites

BACKGROUNDPorcine liver performing efficient physiological functions in the human body is prerequisite for successful liver xenotransplantation. However, the protein differences between pig and human remain largely unexplored. Therefore, we investigated the liver expression profile of a highly inbred minipig line.

METHODSA cDNA library was constructed from liver tissue of an inbred Banna minipig. Two hundred randomly selected clones were sequenced then analysed by BLAST programme.

RESULTSAlignments of the sequences showed 44% encoded previously known porcine genes. Among the 56% unknown genes, sequences of 72 clones had high similarities with known genes of other species and the similarities to human were mostly above 0.80. The other 40 clones showing no similarity to genes in National Centre for Biotechnology Information are newly discovered, expressed sequence tags specific to liver of inbred Banna minipig. Twenty-two of the 200 clones had full length encoding regions, 38 complete 5' terminal sequences and 140 complete 3' terminal sequences.

CONCLUSIONThese newly discovered expression sequences may be an important resource for research involving physiological characteristics and medical usage of inbred pigs and contribute to matching studies in xenotransplantation.

Animals ; Expressed Sequence Tags ; Gene Library ; Liver ; metabolism ; Sequence Alignment ; Swine ; Swine, Miniature ; Transplantation, Heterologous

Rhesus monkeys (Macaca mulatta) are human's closest evolutionary relatives next to Chimpanzees, and they are widely used in biomedical researches. Analyses of the rhesus monkey trasnscriptome and the sequence divergence between monkey and human are of importantce to the development of scientific analyses and to the application and interpretation of the results from animal experiments. In this study, we analyzed the genetic and transcriptional information. Four hundred and one clones were randomly selected from a liver tissue cDNA library of rhesus monkey, and the expressed sequence tags (ESTs) were sequenced. We acquired 393 effective ESTs that were assembled into 221 Unigenes with Phrap software. Alignments of the sequences showed that 188 Unigenes matched with known proteins in Swiss_prot database, of which 16 Unigenes matched the known rhesus proteins, and 171 Unigenes had high homology with human proteins. Then the result of BLASTN comparison showed that 26 of another 33 Unigenes matched the known rhesus genes. Finally, the remaining Unigenes were aligned in dbEST and rhesus genome database, and the results suggested 3 Unigenes be newly discovered ESTs of rhesus.
Animals ; Expressed Sequence Tags ; chemistry ; Gene Library ; Liver ; chemistry ; Macaca mulatta ; genetics ; Sequence Analysis, DNA

Four hundred and sixty five randomly selected clones from a cDNA library of Blattella germanica were partially sequenced and searched using BLAST as a means of analyzing the transcribed sequences of its genome. A total of 363 expressed sequence tags (ESTs) were generated from 465 clones after editing and trimming the vector and ambiguous sequences. About 42% (154/363) of these clones showed significant homology with other data base registered genes. These new B. germanica genes constituted a broad range of transcripts distributed among ribosomal proteins, energy metabolism, allergens, proteases, protease inhibitors, enzymes, translation, cell signaling pathways, and proteins of unknown function. Eighty clones were not well-matched by database searches, and these represent new B. germanica-specific ESTs. Some genes which drew our attention are discussed. The information obtained increases our understanding of the B. germanica genome.
Sequence Alignment ; Reverse Transcriptase Polymerase Chain Reaction ; Molecular Sequence Data ; Male ; Female ; *Expressed Sequence Tags ; Blattellidae/*genetics ; Base Sequence ; Animals

The EST Knowledge Integrated System, EKIS (http://ekis.kribb.re.kr), was established as a part of Korea's Ministry of Education, Science and Technology initiative for genome sequencing and application research of the biological model organisms (GEAR) project. The goals of the EKIS are to collect EST information from GEAR projects and make an integrated database to provide transcriptomic and metabolomic information for biological scientists. The EKIS constitutes five independent categories and several retrieval systems in each category for incorporating massive EST data from high-throughput sequencing of 65 different species. Through the EKIS database, scientists can freely access information including BLAST functional annotation as well as Genechip and pathway information for KEGG. By integrating complex data into a framework of existing EST knowledge information, the EKIS provides new insights into specialized metabolic pathway information for an applied industrial material.
Data Mining ; Expressed Sequence Tags ; Genome ; Metabolic Networks and Pathways ; Metabolomics ; Models, Biological

To accelerate the molecular analysis of specifically induced antibacterial peptide against pathogen (E. coli), cDNA library prepared from the larvae fatbody of Bombyx mori was examined by the expressed sequence tag (EST) analysis. In a total of 722 clones, 653 clones were unique genes. Of 653 unique genes, 43.2% (282/653 ESTs) was identified as characterized genes, 38.1% (249/653 ESTs) as uncharacterized genes, and 18.7% (122/653 ESTs) as novel ESTs. According to the functional categorization of the characterized genes, 36.2% (102/282 ESTs) was antibacterial proteins. The highest expressed peptides, 78.4% of all the expressed antibacterial proteins (80/102 ESTs), belonged to the cecropin family. The antibacterial effect of selected clones representing novel ESTs based on a phylogenetic analysis was examined against various bacterial strains. None of the clones showed significant inhibitory effect to the bacteria tested. These results suggested that most of the novel molecules induced by E. coli may not act as immune-induced antibacterial peptides in the fatbody.
Bacteria ; Bombyx* ; Clone Cells ; Expressed Sequence Tags* ; Gene Library ; Humans ; Larva ; Peptides

OBJECTIVETo explore the complexity of mRNA in the ejaculated sperm from healthy fertile men.

METHODSSemen samples were collected from 10 healthy fathers. The swim-up method was adopted to purify the sperm from possible contamination of somatic cells and the spermatozoal total RNA extracted by Trizol was used for SAGE library analysis.

RESULTSA totle of 21 052 SAGE raw tags were sequenced from 877 clones and 2 712 unique tags that occurred at least twice in the library were given further analysis. 19.7% of the unique tags had no match in the existing SAGE map, representing novel genes. Molecular function analysis revealed 67% of unique tags related to protein binding or nucleic acid binding categories, 41% to catalytic activity, 13% to message transducer activity, and 10% to transporter, structural and transcription regulator activity, respectively.

CONCLUSIONThere exists a complex repertoire of mRNAs in the ejaculated spermatozoa from fertile men.

Adult ; Ejaculation ; Expressed Sequence Tags ; Gene Expression Profiling ; Humans ; Male ; RNA, Messenger ; genetics ; Spermatozoa ; physiology