TIGR Bos taurus Gene Index

Definitions:

• TCs:

  Tentative Bos taurus Consensus sequences are created by assembling ESTs into virtual transcripts. In some cases, TCs contain full or partial cDNA sequences (ETs) obtained by classical methods. TCs contain information on the source library and abundance of ESTs and in many cases represent full-length transcripts.

  TCs are actual assemblies, with a consensus sequence, and not simply clusters of overlapping sequences. Example TC590.

• ESTs:

  Expressed Sequence Tags are partial, single-pass sequences from either end of a cDNA clone. The EST strategy was developed to allow rapid identification of expressed genes by sequence analysis.

• ETs:

  TIGR's Expressed Gene Anatomy Database (EGAD) contains a non-redundant set of nucleotide sequences that represent mature Bos taurus transcripts (ETs). The ETs are curated for nomenclature and cellular roles, and links have been made to related accessions. Sequences were either loaded directly from GenBank (cDNAs) or were derived from genomic sequences when cDNAs were not available. Where available, 5' and 3' non-coding regions were included. Alternative splice forms of genes are explicitly represented.

• Singleton ESTs:

  Also refered to as singletons, are ESTs that are not contained in an assembly. These ESTs went through the assembly process but did not meet the match criteria (see below) to be assembled with any other EST in the collection of ESTs and other GenBank sequences used to create the consensus sequences for a particular Gene Index.

Protocol for Assembly of ESTs and Transcripts

Preparation of EST data

• Sequences were extracted from dbEST and were subjected to quality control screening (vector, polyA, T, or CT removal, minimum length = 100 bp, < 3% N).

Preparation of non-redundant Bos taurus transcript (ET) database

• All Bos taurus sequences from GenBank were extracted.
• Non-coding sequences were discarded and cDNAs and coding sequences from genomic entries were saved.
• Redundant entries for the same gene were removed, retaining link to accession number.
• Sequences and related information are stored in TIGR's Expressed Gene Anatomy Database (EGAD).
• The curated ET data set is available as a multiple FastA format file. Please see the EGAD main page for more information.

Assembly

• Cleaned EST sequences and non-redundant transcript (ET) sequences were combined.
• Using the CAP3 Sequence Assembly Program sequences were assembled into contigs. TCs are consensus sequences based on two or more ESTs (and possibly an ET) that overlap for at least 40 bases with at least 95% sequence identity. These strict criteria help minimize the creation of chimeric contigs. These contigs are assigned a TC (Tentative Bos taurus Consensus) number. TCs may comprise ESTs derived from different tissues.
• The best hits for TC's were assigned by searching the TC set against a non-redundant amino acid database(nraa) using a search method developed in house called DNA-Protein Search, DPS (Microbial and Comparative Genomics, Vol 1, Number 4 1996). The top five hits based on score (cutoff value of 350) were selected and displayed for each TC.

• TCs are only as good as the ESTs underlying them; there may be chimeric ESTs and thus TCs
• There is still redundancy in the TC set because sequences must match end to end and at a certain percent identity to be combined
• Directionality of the TCs should not be assumed
• Not all TCs contain protein-coding regions

References related to EST strategy [Entrez links]

Adams, MD et. al., "Sequence identification of 2,375 human brain genes", Nature 355: 632-4 (1992) [92168112]

Adams, MD et. al., "3,400 new expressed sequence tags identify diversity of transcripts in human brain", Nat Genet 4, 256-67 (1993) [93364420]

Adams, MD et. al., "Rapid cDNA sequencing (expressed sequence tags) from a directionally cloned human infant brain cDNA library", Nat Genet 4, 373-80 (1993) [94004965]

Adams, MD et. al., "Initial assessment of human gene diversity and expression patterns based upon 83 million nucleotides of cDNA sequence", Nature 377(Suppl.): 3-174 (1995) [96026280].