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elg1, rad24 and ctf7 single mutant cells all exhibited relatively little growth defects upon exposure to hydroxyurea.
Upon release from G1, wildtype, rad61 and ctf18 single mutant cells and rad61 ctf18 double mutant cells and also ctf7 rad61 ctripleriple mutant cells all exited S-phase in synchrony such that the time interval from G1 to mid-replication and then to G2/M accumulation is nearly identical (Figure 4).
In light of this partial rescue of proliferation of Vhl mutant cells, all subsequent proliferation experiments were conducted at 5% oxygen in a conventional incubator.
The delay is largely eliminated in ndc80 -K204E ipl1 -2 mutant cells; all cells returned to G1 by the 180-minute time point.
Similarly, in chimeras with a low contribution of mutant cells, all Pax6 −/− progenitors were Gsx2−, even those in very small isolated groups, whereas all wild-type progenitors, even individually isolated ones, were Gsx2+.
In Figure 6D, the colonies derived from added doubly mutant cells all appeared on day 2, while the revertant colonies derived from the tester cell population appeared later and presumably arose from pre-existing singly mutant cells in the tester population that required time and a second mutation before they could make a visible colony.
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Three genes (sdaA, sdaB, tdcG) code for three l-serine deaminases in E. coli, and participate in glycine, serine, and threonine metabolism and cysteine metabolism, and null mutant cells for all three genes grow normally in glucose minimal medium.
MEF 3T3, wildtype, FAK and SYF (src/fyn/yes triple mutant) cells were all maintained in Dulbecco's modified MEM DMEMM, Gibco) supplemented with 10% fetal bovine serum (FBS, Gibco), 2 mM L-glutamine (Gibco) and penicillin G (200 U ml−1, Gibco) in a humidified 37°C incubator at 5% CO2.
This eliminated lawn cells and assessed the aggregate number of mutant cells (in all colonies together) present at each time point.
We performed this dynein composition analysis twice for dyf13 vs wild-type and five times for dyf13oda1 vs oda1 mutant cells, in all cases obtaining similar results.
The α5 integrin subunit was more abundant in these cells than in control GFP-transfected cells or in GFP::mATX3C14 (DUB catalytic mutant) overexpressing cells (all presenting the same levels of endogenous mATX3) (Figure 6A, B), indicating that mATX3 is able to stabilize this protein and prolong its half-life.
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