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To this end, maximum transcript alignments [ 52] of corresponding Unigene clusters [ 53] were generated using SPA [ 54] and examined for instances of: i) introns more than 50 nt downstream of the termination codon of the largest ORF within a transcript, as an indication of AS-NMD.
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The valid EST alignments and cluster alignments were then grouped based on genome mapping location and assembled into gene structures that include the maximum number of compatible transcript alignments.
Errors identified by transcript alignments are generally of four types.
For cross-species transcript alignments, the identity cutoff was decreased to 80%, which selects reliable alignments from divergent transcript sequences.
These two along with PASA transcript alignments were used as transcript and protein evidences for EVM.
We used GenomeThreader [ 61] transcript alignments in exchange of PASA's standard GMAP alignments [ 62] as GenomeThreader supports alignment of transcript evidence to multiple (nearly identical) loci.
GLEAN uses transcript alignments to support splice predictions, but does not merge or split genes based on transcript alignments, while MAKER2 creates gene models that best agree with transcript alignments, and works best with highly reliable transcript assemblies.
The mean transcript length, maximum transcript length, and N50 size of Trinity transcripts were 1112, 22,887, and 1863, respectively.
We also assembled 369,092 spliced Arabidopsis transcript alignments into 29,183 alignment assemblies, of which 22,951 include FL-cDNAs (Table 1).
Transcript alignments with less than 90% EST length coverage or less than 95% alignment identity were discarded.
The PASA-filtered transcripts were then anchored to the genome scaffolds by assembling overlapped transcript alignments into maximal alignment assemblies in order to remove multiple spliced isoforms.
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Justyna Jupowicz-Kozak
CEO of Professional Science Editing for Scientists @ prosciediting.com