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The results indicate that C. reinhardtii transformants were able to produce 63 and 94% more neutral lipids than the wild-type, which translates to an approximately 56% improvement in total lipids, without compromising growth.
Following isolation and transfer of hygromycin-resistant mycelial slices to media containing increasing concentrations of hygromycin B, we found that transformants were able to propagate in the presence of hygromycin B concentrations as high as 300 μg/ml (Figure 3c, bottom panel).
All of the transformants were able to grow on YM plates containing 4.5 mM 5-FOA, indicating that the URA3 reporter gene in the prey plasmid was not expressed.
We obtained approximately 36000 independent transformants of which approximately 500 transformants were able to grow on the galactose-containing plates.
All the transformants were able to grow on SC-His, demonstrating the presence of the expression vector or target gene.
The transformants were able to grow in the presence of the antibiotic paromomycin and produced a detectable level of the AphVIII protein.
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This indicates that transformants are able to express and secrete laccase3 into medium and that the expressed recombinant laccase3 maintains biochemical characteristics of oxidizing a phenolic compound such as tannic acid.
The transformants that were able to grow on resistant plates were considered genetically stable.
In both transformants that were able to grow on glucose medium, PFK1 activity was detected in the homogenates.
To this end, genomic DNA was isolated from transformants that were able to grow for three weeks under selective pressure by exposure to paromomycin.
Considering the haa1Δ transformants, the suppressors were able to rescue haa1Δ susceptibility phenotype by shortening the latency period by at least 20 h when compared with the haa1Δ strain transformed with the empty vector.
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