Journal of the American Society for Horticultural Science, 1993
Regeneration from apple (Malus × domestica Borkh.) M.26 leaf tissue was completely inhibited by (... more Regeneration from apple (Malus × domestica Borkh.) M.26 leaf tissue was completely inhibited by (μg·ml-1) 1 geneticin, 5 kanamycin, 10 to 25 paromomycin, and 100 neomycin. nptII-transgenic M.26 had an increased tolerance to all four of the antibiotics tested, with inhibition of regeneration occurring at (μg·ml-l) 2.5 geneticin, 100 kanamycin, 375 paromomycin, and 375 neomycin. Paromomycin (100 to 250 μg·ml-l) and neomycin (250 μg·ml-1) significantly increased the amount of regeneration from nptII-transgenic M.26 apple leaf tissue. p35SGUS-INT, a plasmid with a chimeric b -glucuronidase gene containing a plant intron, was useful for studying the early events of apple transformation by eliminating GUS expression from Agrobacterium tumefaciens. It was used to determine that the optimal aminoglycoside concentrations for the selection of nptII-transgenic M.26 cells were (μg·ml-1) 2.5 to 16 kanamycin, 63 to 100 neomycin, and 25 to 63 paromomycin. Geneticin was unsuitable as a selective ag...
Journal of the American Society for Horticultural Science, 1998
Seven nptII and gus transgenic lines of the apple (Malus ×domestica Borkh.) rootstock Malling 7 (... more Seven nptII and gus transgenic lines of the apple (Malus ×domestica Borkh.) rootstock Malling 7 (M.7) were examined by glucuronidase (GUS) histochemical testing and a double-antibody sandwich enzyme-linked immunosorbent assay (ELISA). These lines had different amounts of neomycin phosphotransferase II (NPTII). The amounts of NPTII among lines was positively correlated with the ability of the transgenic lines to regenerate in the presence of kanamycin, paromomycin, or geneticin. Regenerants derived from transgenic lines also varied greatly in GUS expression. The apical portion of regenerant stem tissues had stronger GUS staining than the basal portion of stem. All regenerated tissue of T1, a transgenic line originally classified as a uniform GUS staining line, showed non-GUS staining, while the regenerated tissues of chimeric transgenic lines showed nonstaining, chimeric staining, or uniform GUS staining, indicating the potential to select uniform GUS staining lines from chimeras. Po...
Journal of the American Society for Horticultural Science, 2002
Genes encoding lysozyme (T4L) from T4 bacteriophage and attacin E (attE) from Hyalophora cecropia... more Genes encoding lysozyme (T4L) from T4 bacteriophage and attacin E (attE) from Hyalophora cecropia were used, either singly or in combination, to construct plant binary vectors, pLDB15, p35SAMVT4, and pPin2Att35SAMVT4, respectively, for Agrobacterium-mediated transformation of `Galaxy' apple, to enhance resistance to Erwinia amylovora. In these plasmids, the T4L gene was controlled by the cauliflower mosaic virus 35S promoter with duplicated upstream domain and the untranslated leader sequence of alfalfa mosaic virus RNA 4, and the attE gene was controlled by the potato proteinase inhibitor II (Pin2) promoter. All transgenic lines were screened by polymerase chain reaction (PCR) for T4L and attE genes, and a double-antibody sandwich enzyme-linked immunosorbent assay for neomycin phosphotransferase II. Amplification of T4L and attE genes was observed in reverse transcriptase-PCR, indicating that these genes were transcribed in all tested transgenic lines containing each gene. The ...
A chemically inducible and a light inducible DNA promoter are being developed and evaluated for t... more A chemically inducible and a light inducible DNA promoter are being developed and evaluated for their ability to regulate gene expression in transgenic apple. The estradiol-induced XVE gene expression system was cloned into a binary vector (pBinPlusARS) compatible for use in Agrobacterium-mediated transformation of apple. To evaluate the estradiol-inducible binary vectors, a GUS marker gene was cloned into pBinPlusARS.XVE and pBinPlusARS.XVE.Gateway. When evaluated in tobacco under non-inducing conditions, GUS expression from the GUS containing XVE constructs was indistinguishable from that of an empty vector (no GUS coding region) negative control. However, in the presence of estradiol, GUS expression from the GUS containing XVE constructs was greater than that from a 35S promoter positive control. To evaluate the activity of the light inducible promoter of the peach chlorophyll a/b binding (CAB) protein and XVE in transgenic apple, binary vectors were constructed to allow comparison of test promoter activity with 35S activity.
Journal of the American Society for Horticultural Science, 1993
Regeneration from apple (Malus × domestica Borkh.) M.26 leaf tissue was completely inhibited by (... more Regeneration from apple (Malus × domestica Borkh.) M.26 leaf tissue was completely inhibited by (μg·ml-1) 1 geneticin, 5 kanamycin, 10 to 25 paromomycin, and 100 neomycin. nptII-transgenic M.26 had an increased tolerance to all four of the antibiotics tested, with inhibition of regeneration occurring at (μg·ml-l) 2.5 geneticin, 100 kanamycin, 375 paromomycin, and 375 neomycin. Paromomycin (100 to 250 μg·ml-l) and neomycin (250 μg·ml-1) significantly increased the amount of regeneration from nptII-transgenic M.26 apple leaf tissue. p35SGUS-INT, a plasmid with a chimeric b -glucuronidase gene containing a plant intron, was useful for studying the early events of apple transformation by eliminating GUS expression from Agrobacterium tumefaciens. It was used to determine that the optimal aminoglycoside concentrations for the selection of nptII-transgenic M.26 cells were (μg·ml-1) 2.5 to 16 kanamycin, 63 to 100 neomycin, and 25 to 63 paromomycin. Geneticin was unsuitable as a selective ag...
Journal of the American Society for Horticultural Science, 1998
Seven nptII and gus transgenic lines of the apple (Malus ×domestica Borkh.) rootstock Malling 7 (... more Seven nptII and gus transgenic lines of the apple (Malus ×domestica Borkh.) rootstock Malling 7 (M.7) were examined by glucuronidase (GUS) histochemical testing and a double-antibody sandwich enzyme-linked immunosorbent assay (ELISA). These lines had different amounts of neomycin phosphotransferase II (NPTII). The amounts of NPTII among lines was positively correlated with the ability of the transgenic lines to regenerate in the presence of kanamycin, paromomycin, or geneticin. Regenerants derived from transgenic lines also varied greatly in GUS expression. The apical portion of regenerant stem tissues had stronger GUS staining than the basal portion of stem. All regenerated tissue of T1, a transgenic line originally classified as a uniform GUS staining line, showed non-GUS staining, while the regenerated tissues of chimeric transgenic lines showed nonstaining, chimeric staining, or uniform GUS staining, indicating the potential to select uniform GUS staining lines from chimeras. Po...
Journal of the American Society for Horticultural Science, 2002
Genes encoding lysozyme (T4L) from T4 bacteriophage and attacin E (attE) from Hyalophora cecropia... more Genes encoding lysozyme (T4L) from T4 bacteriophage and attacin E (attE) from Hyalophora cecropia were used, either singly or in combination, to construct plant binary vectors, pLDB15, p35SAMVT4, and pPin2Att35SAMVT4, respectively, for Agrobacterium-mediated transformation of `Galaxy' apple, to enhance resistance to Erwinia amylovora. In these plasmids, the T4L gene was controlled by the cauliflower mosaic virus 35S promoter with duplicated upstream domain and the untranslated leader sequence of alfalfa mosaic virus RNA 4, and the attE gene was controlled by the potato proteinase inhibitor II (Pin2) promoter. All transgenic lines were screened by polymerase chain reaction (PCR) for T4L and attE genes, and a double-antibody sandwich enzyme-linked immunosorbent assay for neomycin phosphotransferase II. Amplification of T4L and attE genes was observed in reverse transcriptase-PCR, indicating that these genes were transcribed in all tested transgenic lines containing each gene. The ...
A chemically inducible and a light inducible DNA promoter are being developed and evaluated for t... more A chemically inducible and a light inducible DNA promoter are being developed and evaluated for their ability to regulate gene expression in transgenic apple. The estradiol-induced XVE gene expression system was cloned into a binary vector (pBinPlusARS) compatible for use in Agrobacterium-mediated transformation of apple. To evaluate the estradiol-inducible binary vectors, a GUS marker gene was cloned into pBinPlusARS.XVE and pBinPlusARS.XVE.Gateway. When evaluated in tobacco under non-inducing conditions, GUS expression from the GUS containing XVE constructs was indistinguishable from that of an empty vector (no GUS coding region) negative control. However, in the presence of estradiol, GUS expression from the GUS containing XVE constructs was greater than that from a 35S promoter positive control. To evaluate the activity of the light inducible promoter of the peach chlorophyll a/b binding (CAB) protein and XVE in transgenic apple, binary vectors were constructed to allow comparison of test promoter activity with 35S activity.
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Papers by Herb Aldwinckle