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4.155. Molecular analyses of transgenic plants
4.15.5.1. PCR, Southern and Western blot analyses of call tronsfor~nonts
When the genomic DNA of cyanamide tolerant tr:~nsforni:mts was uhcd :IS
template, gene-specific primers amplified
7 3 2
hp of coh gene in the 6 transfor~~i;~nts
and positive control (pCAMBIA-cah). Conversely, no such hand was observed in the
untransformed control under identical conditions (Fig 3 IA).
Southern blot analysis was carried out on a11 the tronsfur~uants using cah
coding sequences as probes. Two transformants (plant I and 3 ) displayed one
hybridizable band and 4 transformants (plant
2 , 4 .
5 and 6) exhibited two hyhridizable
bands for rah probe from the total DNA of transformants indicating the slahle
integration of marker gene rah in transgenic tobacco plants (Fig. 31B).
Transgenic tobacco plants were studied for the expression of cyanamide
hydratase. Cyanamide hydratase antisemm detected expression of
27.7
kDa
polypeptide in all 6 transformants, whereas such expression was absent in the
unuansfonned control plant (Fig. 3 1C).
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