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To follow var-gene switching over time, two parasite clones were monitored for ≈200 generations during which, both RNA expression and phenotypic changes were investigated at least once a month.
Clones were monitored for RFP expression by fluorescence microscopy (Leica DMIRB microscope with appropriate filter sets) before isolation and one to three times per week after isolation.
Then, the levels of CXCL12, CXCR4, and CXCR7 transcripts in the control clones and COUP clones were monitored using real-time quantitative RT-PCR.
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Growth of HeLa clones was monitored by fluorescence staining of the DNA with Hoechst 33258 (Sigma).
To ascertain whether the lack of interaction between MP and P8, RdRp or Pro domains was not due to the lack of expression of the proteins in the yeast cells, the expression of all the proteins, both from pGAD T7 and pGBK T7 clones was monitored using HA polyclonal and cMyc monoclonal antibodies respectively using the lysed transformed AH109 cells grown in -Leu -Trp -His medium.
(B ) Growth of the TbHpHbR null clones was monitored in vitro over 192 hr.
To assess the susceptibility of bacterial populations to infection by phages, bacterial growth of isolated bacterial clones was monitored as optical density with and without phages.
Possible loss of the VAP in single clones was monitored regularly by immunodot blot using a monoclonal virulence-associated protein A (VapA) antibody (10G5; Takai et al., 1993).
Subsequently, 7.5 μl of culture were dropped on vector- and interaction-selective media (CSM-L-, W-, Ade-) and incubated at 28 C. At day 2 the growth of the clones was monitored.
For expression analysis in living cells, the published promoter regions, or putative regulatory 5'FR of up to 2 Kb, were cloned and basal levels of luciferase were monitored.
However, an abnormal aneuploid clone was not detected as far as the cells were monitored by a DNA histogram analysis.
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