To breed A. aculeatus that simultaneously produces many types of cellulases, a marker-recycling technique, enabling to excise a selection marker gene from transformants without leaving any exogenous DNA fragments, must be developed (Akada et al. [2006]).
Acylation activities in the LPEAT transformants without addition of any lysophospholipid acceptor indicated there were endogenous lysophospholipids in microsomal extracts [see Additional file 3].
Two recent articles report different and novel strategies to either remove antibiotic resistance genes or select chloroplast transformants without using these genes.
As shown in Fig. 4C, although hTERT overexpression reduced BMP7-induced cell death significantly, BMP7 treatment of GFP-hTERT transformants still showed more cell death than the GFP-hTERT transformants without BMP7 treatment (P = 0.001) (Fig. 4C).
This technique enables the excision of a selection marker gene from transformants without leaving exogenous DNA fragments; multiple cellulase genes can be introduced into a host strain with a limited number of selection markers.
The availability of these constructs made it possible to generate an approximately uniform mixture of corresponding transformants without induction, and then to test the impact of their expression when galactose was added.
The expression of the luciferase was analyzed in detail after a long-term incubation of the transformants without selective pressure (see below).
At the gradient annealing temperatures from 64 ~ 69°C, the WT template, as well as two clones (D112K-1 and D112K-3) oft ofiveve randomly collected from the transformants without Dpn I digestion, showed detectable products only at annealing temperatures of 64 and 65°C, whereas the other three clones (D112K-2, D112K-4 and D112K-5) maintained detectable up to 67°C or higher.
We have found the HpGSH2 gene overexpression in mcGSH2 transformant cultivated in medium without any carbon substrate.
Analysis of pools of transformants grown without induction already identified some differential depletion of cDNAs, presumably because unspecified factors were sequestered by the inserts.
