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To avoid false positive due to potential pleiotropic effects of the genetic markers, we performed a leave-one-out sensitivity analysis and an MR-Egger sensitivity analysis that we expanded in the survival context.
To provide a more comprehensive view of Pokkali QTLs using SSR markers, we performed a new QTL study using 100 genome-wide SSRs on 140 RILs, including 64 of the extreme RILs plus 76 randomly selected lines from the original population of 276 RILs developed from the cross of Pokkali x IR29 (Gregorio 1997).
Then, using a panel of carbohydrate surface markers, we performed flow cytometry analysis.
To verify that these ERα positive AVPV neurons actually express VGAT and VGLUT2 markers, we performed confocal microscopy on brain sections triple labeled for VGAT, VGLUT2 and ERα markers.
In addition to the analysis of candidate gene-associated markers, we performed a genome-wide association study to validate linkage to LAMC2 and determine if additional genomic regions displayed evidence of association.
Since multiple cluster 1 measures and UAW were found to be associated with similar sets of SNP markers, we performed a principle component analysis (PCA) to evaluate whether some "lower-dimensional summary" of these variables provide more information about the observed associations.
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To select specific CpG sites that would be most promising as a noninvasive fetal DNA marker, we performed bisulfite sequencing on the 4 selected regions.
As c-kit is a differentiating spermatogonia marker, we performed qPCR and western blot assays to detect its transcription and translation activities.
As a third independent marker, we performed a combined analysis of two regions of the mitochondrial genome: since haplowebs are not suited to analyze non-recombining markers, individuals were sorted into four haplogroups according to their mitochondrial sequences.
To assess whether decreased invasion observed in vitro and in vivo is associated with changes in EMT markers expression, we performed qPCR to assess the transcript levels of EMT markers genes SNAI1 and ZEB2.
On this marker set, we performed single-marker transmission disequilibrium tests (TDTs) [37] using PLINK v1.07.
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