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The maximum sequence coverage was obtained from G-1186/94 followed by Atlas with 16X and 15X, respectively (Table 1).
To achieve maximum sequence coverage of the protein we combined proteolytic digests from four proteolytic enzymes (trypsin, chymotrypsin, LysC, and subtilisin).
Results have implications for future analyses of membrane proteins when maximum sequence coverage is required (e.g., mapping protein protein interactions, identification of isoforms, and post-translational modifications), and moreover, for quantitation of membrane and soluble proteins when reproducible and specific enzymatic digestion is required.
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To maximize assembly efficiency, a minimum of 25x and maximum 400x sequence coverage at RAD SE reads were imposed.
To maximize efficient assembly of sequences we imposed a minimum of 50x and maximum 750x sequence coverage at any RAD sequence cluster.
All the hits of the earlier approaches were coupled with manual intervention to ensure maximum residue and sequence coverage of Mtb proteome (29).
For peptide matching, a minimum of four peptides matches and 15% sequence coverage, a maximum of one miscleavage, and peptide modifications by carboxyamidomethylcysteine, methionine sulfoxide, and pyro-glutamic acid or acetylated N-terminal residue, were accepted.
Similarly, on average 64% of the maximum sequence length improvements were seen with 25× coverage, and 60% of the N50 gains.
The consensus sequence obtained from this alignment showed significant homology to gamma-TMT haplotypes 1, 3, 4 and 5 (GenBank accession nos. DQ229828, DQ229830, and DQ229831 and DQ229834; 8e−18), with 89% maximum sequence identity spanning 18% of the fragment coverage.
The maximum sequence reads for single contig is 505,778, with the coverage of 62,678.
The consensus sequence for locus c (388 bp long) showed a significant homology to gamma-TMT haplotypes 4 and 5 (GenBank accessions nos. DQ229831 to DQ229834, 4e-14), with 86% maximum sequence identity spanning 19% of the locus c fragment coverage.
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