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(E) A ssDNA oligo (Oligo donor) encoding 6 silent mutations (indicated in red) was synthesized (top), and co-transfected with the G1 gRNA-Cas9 construct pX330-G11) into 293T cells (middle).
Drosophila Fbxl7 mutations indicated in red.
Point mutations indicated in red showed reduced binding affinity to Usp.
(D ) Cartoon of IGF1R TM-icd with the positions of single-site cysteine mutations indicated in red (top).
AR + cell lines (indicated in green, Figure 3) and other TNBC cell lines containing PIK3CA mutations (indicated in red, Figure 3) were the most sensitive to pharmacological PI3K inhibition, as indicated by low, half-maximal inhibitory concentration (EC50) values.
Human WDR73 (NM_032856.2) and CAD (NM_004341.3) cDNAs amplified from HEK-293T total RNA using RT-PCR (SuperScript® III reverse transcriptase, Life Technologies Phusion DNAA polymerase, NEB) were cloned into pENTR/D-TOPO (Life Technologies) and mutations indicated in the 'Results' section were introduced by site-directed mutagenesis (QuikChange® II, Agilent).
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However, separate analysis of the samples from women bearing BRCA1 mutations indicated that, in the absence of E2, the epithelial Ki67LI decreased significantly to a median of 0.7% (IQR=0.4 1.0; n=21; P=0.009 by Mann–Whitney U-test).
cdc13-1 strains with additional mutations indicated were grown in YEPD at 23 °C overnight.
The sequence context of C mutations in unclustered and kataegic locations are shown (lower plots), with the number of mutations indicated, coloured as in Figure 3B.
The spectrum of these mutations is indicated in Figure 2 and Table 2. Sixty percent of the mutations occurred at three sites bp 172 (R58X), bp 238 (R80X) and bp 247 (H83Y)–and were all C→T transitions.
Primer sequences for amplifying FGFR2, deleting 1 kbp in the middle of intron 9, and introducing mutations were indicated in Methods S1.
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