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The method proposed here is based on identifying the recombinant predictions which are supported by the three methods.
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In order to identify the recombinant proteins, V5, Flag and GFP tags were added to the C-termini (Fig 1A).
A structural analysis identified the recombinant molecule as predominantly disordered.
The Flag epitope identifies the recombinant protein but not the endogenous form of SIRT-8.
SDS-PAGE and Western blot analysis were applied to analyze the purity and to identify the recombinant swollenin.
However, only one marker RM80, flanking the gene xa13 at a distance of 1.1 Mb, was found to be polymorphic and this was used to identify the recombinant line with minimum donor parent segment (Fig. 1A).
To identify the recombinants with least donor segment introgression, 22 markers were identified from the flanking regions (∼5 Mb on either side) of the gene xa13 (25.01 Mb) on chromosome 8.
Accordingly, specific substrates for major enzymes of GH43, like α-l-arabinofuranosidases (AFs), and endo-α-l-arabinanase (endo-ABNs), exo-α-l-arabinanases (exo-ABNs), and so on were chosen to measure the enzyme activity for identifying the function of recombinant protein.
Recombinant viruses (ECTVΔCD30, ECTVRevCD30 and ECTVmCD30) were finally selected by successive plaque purification of white plaques in the absence of puromycin and screening by a CD30-specific PCR to identify the desired recombinant viruses.
It uses the familiar blue-white screening technique to identify the desired recombinant clones.
Using complete genome sequence analysis, we recently identified the first recombinant (intragenogroup) SaV strains (3 ).
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