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These recoded synthetic drug-resistant genes could be a useful tool for selecting mitochondrial genetic transformants as a precise technology for mitochondrial transformation is developed.
The data presented here illustrate the need to evaluate and characterise genetic transformants thoroughly, since genetic engineering procedures sometimes yield unexpected outcomes.
It is worth mentioning that variation of polyamine levels in the genetic transformants overexpressing polyamine biosynthetic genes has been reported in earlier studies [ 32, 40].
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Genetic transformation of KU-41 protoplasts with 10 µg of the marker plasmid pAH yielded 26 35 transformants; no clones were obtained in the absence of pAH (Table 1).
Further, the level of resistance to antibiotic (2 mg/ml) revealed by different transformants amply testify the integration of 4-7 cofieSAM2 SAM2 gene as these two genes are linked in the recombinant plasmid employed for genetic transformation.
Procedures for S. gordonii transformation, screening and genetic analysis of transformants were as previously described [ 33].
Fig. 4 Assessment of genetic stability of transformants in the absence of selective pressure.
Transformation of naturally competent cells of S. gordonii V288 and GP201, scoring and genetic analysis of transformants was carried out as already described [ 21, 22].
The variation observed resulted from fermentation conditions rather than differences in the genetic background of the transformants.
All archaeal strains used in this work, including the genetic host and its transformants of diverse plasmids as well as the orc1/ whip, are summarized in Table S1.
See transformant; transformation efficiency.
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