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rVISTA 2.0 searches for conserved TFBSs were performed against 500 vertebrate TF matrices from the TRANSFAC library, with matrix similarity cuttoff 0.85 by submitting a BLASTZ alignment file for each CNE to the rVISTA 2.0 site.
Using custom Perl scripts we extracted the portions of the pair-wise genome alignments covered by annotated Release 5 D. melanogaster transcripts (in case of multiple isoforms we selected the longest transcript to represent the gene) and collected the pair-wise alignment into single 'multiple' alignment file for each gene.
Sigma generates a BAM alignment file for a selected reference genome using only its assigned reads.
The alignment file for the base element family was used to determine if different parts of the base element made up a complete element.
Additionally, the seed alignment file for the Hsf domain (PF00447) obtained from the Pfam database [ 72] was used to build a HMM file using the HMMER3 software package [ 73].
In order to produce the input alignment file for QRNA v.2.0.3.c [ 7], the 61 ncRNAs annotated in the E. coli K12 Refseq file (NC_000913) were extracted, and compared to the chromosomal IGRs of S. meliloti thanks to WU BLAST 2.0, with various e-value thresholds (10-5, 10.1, 0.1 and default: 10).
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The FASTA alignment files for each of the genes were imported into MacClade 4.08 [26] for manual editing.
The MacClade alignment files for each of the 37 genes can be obtained from authors upon request.
The resulting multispecies alignment files for each analyzed gene are available at http://giladlab.uchicago.edu/data.html.html
Raw count values were determined using HTSeq count [ 63] (−m union) using the generated GFF file and individual SAM alignment files for each sample.
Phylogenetic trees are imported from Ensembl and users can also access the gene tree and alignment files for the specific RBP being queried.
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