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For genotyping, about 3 5 mm of tail tip (soft tissue only) was clipped using a sterile, sharp scalpel.
Approximately 2 3 mm of tail clipping from individual neonates was placed into a 1.5 ml microfuge tube and DNA extraction was carried out using the High Pure PCR Template Preparation Kit (Roche) following the manufacturers protocol.
Each spring, we capture Anolis sagrei lizards from the large island of Great Exuma, sample a small piece (2 mm) of tail tissue from each individual for genetic analyses, and introduce propagules of ca. 200 lizards to each of several offshore islands which we have previously denuded of resident lizards.
For every centimetre SVL growth gained by an aquatic salamander, it gained approximately 30 mm of tail height; while every centimetre of snout-vent growth gained by a terrestrial salamander was accompanied by only 12 mm of tail height.
Samples were processed in 15 batches of ∼50 animals each and some failed samples were removed; 5 mm of tail were collected for genotyping.
A total of 1 mm of tail tip was removed using a scalpel blade, and the tail tip was bathed in sterile saline at 37°C.
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For genotyping, 1 2 mm sections of tail tip were dissolved in 0.1 ml of 50 mM Tris (pH 8.0), 100 mM EDTA, 0.5% SDS, and 0.5 mg/ml proteinase K (Roche) solution at 55°C over night with vigorous shaking.
DNA was isolated from ∼2 mm of mouse tail using the Wizard Genomic DNA purification Kit (Promega, Madison, WI) tail tissue protocol.
The scab was removed for subsequent blood sampling, until 2 mm of the tail tip was removed.
Genomic DNA was isolated by incubating rat tail (−3 mm) in 500 µl of tail lysis buffer (100 mM Tris-HCl [pH 8.0], 5 mM EDTA [pH 8.0], 200 mM NaCl, 0.2% [w/v] SDS, and proteinase K 100 µg/ml) overnight at 55°C.
After 5 min, 2 mm of distal mouse tail was removed by scalpel.
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