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After filtering out the sequences mapped to short clusters of length less than 50 bp, 389,038 bisulfite sequences (75% of all reads) were uniquely mapped to the non-repeat-masked human genome (NCBI build 36.1,hg18).
The majority of piRNAs locate in genome as clusters of length 20-100 kb, and it was suggested that piRNAs are originated from long transcripts then subsequently processed into ~28 nt smaller RNAs [ 9, 41].
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The small RNA clusters of lengths ranging from 47 nucleotides to around 7 kb were retrieved from deepBase [ 26] database.
The procedure is repeated merging bifurcating clusters of lengths d1 and d2 and sizes w1 and w2 connected to a stem-branch of length d s into a larger cluster of average length d = (w1 d1 + w2 d2)/(w1 + w2) + d s and average size w = (d1 w1 + d2 w2 + d s )/ d, until the tree is reduced to two clusters connected to the root (d s = 0).
Each fork-structure comprising two terminal branches (clusters) of lengths d1 and d2 and sizes w1 and w2 bifurcating from a stem-branch of length d s is considered in turn.
Each fork-structure is progressively replaced by a corresponding cluster of length d and size w.
However, consider the case when we have two genomes in a cluster, of lengths 1 Mbps and 5 Mbps respectively.
When the ORF length frequency of E. coli K12 genes with coding-sequence lengths >2000 bases were plotted (Figure 5B), two distinct clusters of ORF length distribution emerged: the NED pattern with an ORF maximum of ∼750 bases, similar to that in random DNA; and a second cluster of non-random, non-conforming ORFs ranging from 2000 to several thousand bases.
Within the filtering procedure, the gene-specific scoring plays a crucial role in distinguishing paralagous gene clusters of equal length.
We restricted the test to clusters of a length of at least 1 year and at most 25% of the population without geographic overlap.
Nonetheless, advances in analysis methods for RNAseq, such as the assembly of clusters of full length alternative spliced isoforms [ 28] and models accommodating isoform expression estimates uncertainties [ 61], allow the estimation of differential gene expression for isoforms in our species.
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