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Additional "mate pair" libraries, produced by circularising genomic DNA fragments of between 3 kilobase pairs (kb) and 10 kb in length, were generated and 50 bp was sequenced from both ends in order to assist with genome assembly.
Ly strain type I was found in most (8 [89%]/9), and a larger amplicon of 762 bp was sequenced from IK/23, which shared little sequence homology with the Ly strain.
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For 12S rDNA, approximately 350 bp were sequenced from R. microplus representing 15 strains from America, Africa, Asia and Oceania.
The first is called paired-end sequencing, where ∼100 bp are sequenced from each end of ∼400-bp DNA fragments.
Fragments larger than 500 bp were sequenced from both ends, after which new primers were sequenced from the preliminary leads to fill gaps for both strands.
Approximately 36,000 cDNA clones (average insert size of 800 bp) were sequenced from both directions and after sequence quality assessment a total of 50,248 ESTs were collected.
Short tags (300 400 bp) were sequenced from the 10 kb and 5 kb amplicons (within the cox1 and rrnL genes, respectively) to verify their specificity and identity.
An additional 9,717 bp were sequenced from portions of thirteen expressed duplicated loci of the tetraploids S. epitropicalis and S. new tetraploid, and 6,966 bp were sequenced from portions of nine expressed duplicated loci of the tetraploids X. muelleri or X. gilli.
The mt genome of the former species was sequenced completely, whereas only about 8,500 bp were sequenced from the mtDNA of Calyptraea chinensis due to unsuccessful PCR amplification of the remaining portion of the genome.
These amplicons were cloned in three of these four species (cloning of this amplicon was unsuccessful in P. bifurca) and approximately 1500 bp were sequenced from each end of the clone insert.
Parts of rnl (~730 bp), a fragment located between Rps3 and the putative N-acetyl-transferase (~850 bp), and a fragment including parts of nad1 and nad4 (~1,650 bp) were sequenced from 32 strains belonging to eight PAC species (Table 3).
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