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A previous attempt to introduce up to 75 Kb of fungal DNA into Fusarium oxysporum and A. awamori using an Agrobacterium tumefaciens transformation system yielded few transformants with large DNA inserts and furthermore, no attempts to examine stability of heterologous DNA, let alone expression, were made [ 34].
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The size of the nanobody library is 8 × 108 transformants with great diversity.
One transformant with the largest halo (P tcu1 - xyr1) was chosen for further analysis.
One transformant with the largest halo GKX21 was chosen for shake-flask fermentation.
In contrast, the pVND7 IRX7 transformants showed large open vessels that were comparable to those of the wild type.
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Strikingly, in all cases, the number of transformants with loss-of-function phenotypes was much larger than those with an overexpression phenotype.
A valuable application of our system beyond studies of morphometric traits could be in the production of transgenic flies, where successful transformants with red eyes have to be sorted from among large numbers of white-eyed flies.
Transformation of the alb1 mutant with pAUR-PksP yielded transformants with black conidia, whereas all transformants of alb1 with pAUR325 remained albino (Figure 3B).
Transformants with elevated laccase production might be generated by modifying the pGL promoter region and/or by screening among a larger number of co-transformants.
As an important prerequisite for a large-scale gene/function correlation study in this plant, we are establishing a collection of Physcomitrella patens transformants with insertion mutations in most expressed genes.
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