Transgenic asparagus (Asparagus officinalis L.) plants were obtained t
hrough particle gun bombardment of suspension cells using the pKGUS pl
asmid containing both neomycin phosphotransferase II (nptII) and P-glu
curonidase (uidA) genes. In experiments to optimize bombardment parame
ters, the combination of 1.1 mu m tungsten particles and 1000 psi bomb
ardment pressure was best; 65 cells or cell clumps per plate of 10(5)-
10(6) suspension culture derived cells expressed uidA in transient ass
ays. No significant genotypic effects were detected for the number of
cells or cell clumps expressing uidA among four genotypes. A two-step
selection procedure was established for the recovery of transformed pl
ants. The bombarded cells were first placed on embryo induction medium
with 25 mg l(-1) kanamycin for 4 weeks, and the resistant calli were
then transferred to the same medium with 50 mg l(-1) kanamycin for emb
ryo development. A total of 10 plants from six kanamycin resistant cal
li were recovered. The expression of uidA was detected in the stems an
d cladodes of these 10 plants. Southern analysis confirmed the integra
tion of nptII and uidA into the asparagus genome, and revealed that th
e 10 transgenic plants from six resistant calli were from one transfor
mation event. In addition to the detection of junction fragments where
the pKGUS plasmid integrated into the genome, Southern analysis also
provided evidence that most of the pKGUS plasmid DNA was integrated in
tactly and organized as head-to-tail concatemers in these plants. uidA
assays of the hybrid seeds from the crosses between one transgenic ma
le asparagus T-0 plant and nontransformed female asparagus suggested t
hat uidA activity was inherited as a single locus with Mendelian segre
gation. (C) 1997 Elsevier Science Ireland Ltd.