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One worm was picked into each well and allowed to equilibrate for at least 5 minutes.
The library was picked into 96 well plates and screened for responsiveness to the addition of the relative effector.
A single colony for each clone was picked into a deep well block containing 1 mL 2xYT, 100 μg/mL ampicillin, 15 μg/mL tetracycline, 0.8% glucose in each well.
In the second approach, individual animals from the strain carrying the mutation of interest were allowed to lay eggs for 2 3 days, and then the parent was picked into 5 μl of lysis buffer and lysed at 57 °C for 1 h followed by 95 °C for 15 m to inactivate the proteinase K.
Transformant E. coli clone (1 discrete colony), confirmed by PCR and sequencing, was picked into 200 mL of 2x YT broth in a 2 Litre flask and shaker-incubated at 35°C for 5 hours to mid-log phase (OD600nM = 0.7).
Similar(55)
White colonies on selective Luria Bertani (LB) agar plates were picked into 96-well blocks containing 1 ml LB broth plus kanamycin (50 µg ml−1) and grown overnight.
Four colonies from each transformation are picked into 96-well plates containing LB medium plus the appropriate antibiotics.
Transformants were picked into 50 ul of ultra-pure water and plasmid DNA liberated by heat lysing.
Plaques were picked into phage buffer and replated with bacteria for several rounds of infection to purify phage isolates.
Plaques were picked into 100 µl phage buffer (10 mM Tris-HCl, pH 7.5; 10 mM MgSO4; 68.5 mM NaCl; 1 mM CaCl2).
A total of ∼16,000 clones were picked into 96-well plates and replicated to 384-well plates for storage at −80°C.
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Justyna Jupowicz-Kozak
CEO of Professional Science Editing for Scientists @ prosciediting.com