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Our dataset contained 230 roo and rooA elements with coding sequences as long as 2615 amino acids, which renders accurate multiple alignment identification challenging.
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In total, multiple alignment and identification of orthologs provided support for the existence of seven polycistronic clusters which concern 38% (15/39) of the Hox genes (table 1).
This preprocessing includes: (i) download of genome sequences and gene annotations, (ii) unification of gene annotations among the set of considered strains, (iii) identification of gene families, (iv) computation of multiple alignments and identification of point mutations which constitute the input genotype data.
On the other hand, PRAC and HyPRE multiple alignments allowed identification of other important and non dissociated elements that account for the discrimination of the enzymes, such as the presence of the aliphatic Cys (or Leu) residue in TcPRAC at position 270 which is absent and replaced by a polar His residue in HyPRE (R2) thus favouring its interaction with OH-Pro.
However, their program relied on prior knowledge of multiple alignments for identification of conserved regions, which may not be generally available for all genomes.
Mugsy was also the fastest tool evaluated for the alignment of four assembled human chromosomes, completing the LCB identification and multiple alignment in <1 h provided a library of pairwise alignments.
The flowchart consists of 4 steps – multiple alignments and consensus identification, information content enhancement, HMMs and sequences logos, and major consensus.
The graph utilized for the LCB identification and segment-based multiple alignment is compact for highly conserved sequences allowing for efficient computation.
This protocol led to the identification of 7985 blocks, representing on average 46% of the total multiple alignment (coverage ranged from <20% to >80%).
For this stage we developed a new R script facilitating the visual identification of both intron position and exon sequence conservation in a multiple alignment.
Structure‐based examination and multiple alignment of sequences revealed SixA active residues from loop β1‐H2, which might assist in the identification of SixA homologs among RHG phosphatases even with poor amino acid identity.
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Justyna Jupowicz-Kozak
CEO of Professional Science Editing for Scientists @ prosciediting.com