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An overabundance of R mutations was considered an indication of positive selection, and a lack of R mutations was considered an indication of negative selection.
Activation of two main EGFR-dependent signaling pathways, the RAS-RAF and the PI3K-PTEN-AKT pathways through mutations was considered to be one of the most common mechanisms involved in colorectal carcinogenesis.
Since tumoral cells in culture may undergo genetic instability the possible presence of DCX deleterious mutations was considered and ruled out by direct sequence analysis (see Materials and Methods) of the SK-N-SH DCX gene coding regions (data not shown).
To detect significant polymorphisms, that is likely polymorphisms per se, we calculated probabilities with the same parameters, save that k = number of deviant reads supporting a particular base at a given position and P = 0.0001/3 because only one of three possible mutations was considered (fig. 1).
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Other mutations are considered to be associated with virulence and clinical severity.
Factor V G1691A (Leiden), prothrombin G20210A, and methylenetetrahydrofolate reductase (MTHFR) C677T mutations are considered risk factors for venous thromboembolism.
BARD1 mutations are considered a high cancer risk and need further examination, as their exact function after mutation is relatively unknown.
Mutations were considered as "not tolerated" when P<0.05.
Intragenic suppressor mutations were considered separate events, and calculated as such.
Introduced mutations were considered to be detectable if their z-score was higher than all of the significance scores for unexpected mutations (false positives) on the control samples.
Mutations were considered to be somatic only if they were found in an independent tumor-derived PCR product but not in matched normal tissue.
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