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In this new HI mutant we sequenced the full genes that were mutated in the original HI (or their surrounding regions, when they were placed in intergenic regions).
To confirm the existence of this mutant, we sequenced the IRF-5 cDNA amplified from a Human T-cell leukemia virus type 1-infected T cell cDNA library and an EBV-infected B cell cDNA library, respectively.
To verify the molecular basis of resistance in the tet mutant, we sequenced the full genome and found only a single point mutation in the α5 region of the MAR repressor marR, which is linked to dimer formation and subsequent binding to the mar promoter [ 33].
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To characterize alterations in gene expression across the whole transcriptome in Adar1 mutant Adar1; Mavs double-mutant embryos, we sequenced ribosomal RNA-depleted total RNA from whole E11.5 embryos.
Each time an assayed mutant was functional, we sequenced the new P450 gene.
Therefore, to increase our likelihood of detecting any developed resistance mutants we performed our sequence analysis on DNA samples from the earliest possible time point at which HIV-1 DNA was detected.
Starting with purified genomic DNAs of strains 34F2 and gerH::ery mutant we obtained the draft sequences of each of these strains in 24 hours using the FLX protocol.
Given both that the phenotype of edr2-6 resembles that of the edr2-1 (a point mutation leading to the change, W256STOP) [ 16] and that the rescue experiment with the cloned EDR2 gene restored the wild-type phenotype to the edr2-6 mutant, we feel that the additional sequences in this construct did not contribute to the phenotypes described in the text.
We sequenced the mutant fab1-2, and identified a single missense mutation, G864E, in the CCT domain.
To examine how frequently this gene is mutated in axenic mutants, we amplified and sequenced it from six further strains: five more new mutants selected from the same parental DdB strain, and one from the V12 genetic background (strains used in this study are listed in Table 2).
Therefore, we sequenced 211 mutants from comfrey-treated rats and 63 mutants from control rats.
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