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Compared with cells transfected with miR-19a/b mimic and control vector, the cells transfected with both miR-19a/b mimic and MTUS1 vector exhibited a significantly lower proliferation rate (Fig. S3A), suggesting that miR-19a/b-resistant MTUS1 can attenuate the proliferative effect of miR-19a/b on lung cancer cells.
Approximately 48 hours after double-transfection with either pDsRed-ph-AKT plus pAcGFP-N1-COMT or pDsRed-ph-AKT plus control vector, the cells were stimulated with NRG1 and terminated by fixation buffer at different time points.
Two days after infection with vector, the cells were lysed with Cell Culture Lysis Buffer (Promega), and luciferase activity was measured as described by the manufacturer.
Following transfection of CHO-K1 cells in a 75 cm flask with this vector, the cells were left undisturbed for 3 weeks, after which substantial numbers of clones were visible by eye.
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Adding the relative risk values makes the assumption that the highest absolute abundance of a vector among the cells of the landscape is the same for each vector.
Two days after vector infection, the cells were lysed, and luciferase activity was measured.
The results showed expression of this protein in all cell lines, ruling out the possibility of complete loss of the vector in the cells derived from LMP tumours.
VT constructed the vectors, generated the cells, performed the induction, radiation and chemotheraphy experiments and writing.
The efficiency of genetic modification of primary cells also depends on the effective introduction of DNA vectors into the cells.
To express shRNA, we transfected shRNA expression vectors into the cells using Lipofectamine 2000 (Invitrogen, Carlsbad, CA).
The high cellular uptake (>80%) and localization of the copolymer vectors inside the cells were easily analyzed by tracking the fluorescence of polythiophene using fluorescent microscopy and cytometry.
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