In this study,cowpea trypsin inhibitor (CpTI) gene, an insecticidal gene,was introduced into two Populus tomentosa clones by gene transformation mediated by Agrobacterium tumefaciens. The transformed regeneration shoo...In this study,cowpea trypsin inhibitor (CpTI) gene, an insecticidal gene,was introduced into two Populus tomentosa clones by gene transformation mediated by Agrobacterium tumefaciens. The transformed regeneration shoots were obtained directly by leaf discs.In order to established selection condition,leaf discs were cultured in the medium with increasing concentration kanamycin.Kanamycin resistant(km r)plantlets were obtained by 3～4 cycles screening in selective condition.Then transformed shoots were rooted in the medium containing kanamycin 50?mg/L and transferred to greenhouse.The presence of CpTI gene in transgenic plants were confirmed by PCR and PCR\|Southern blot.展开更多
Both non-transgenic hybrid triploid poplars [ (Populus tomentosa×P.bolleana)×P.tomentosa ] and transgenic ones expressing cowpea trypsin inhibitor were cut at the base of the stem to produce auxoblasts, an...Both non-transgenic hybrid triploid poplars [ (Populus tomentosa×P.bolleana)×P.tomentosa ] and transgenic ones expressing cowpea trypsin inhibitor were cut at the base of the stem to produce auxoblasts, and used as source of leaves for insect feeding trials performed on 3 major insect species of poplar, the forest tent caterpillar ( Malacosoma disstria L.), gypsy moth ( Lymantria dispar L.) and willow moth ( Stilpnotia candida Staudinger). The height and basal diameter of trees were measured by the end of that year (2000). The results indicated that the growth elements of transgenic poplars were not interfered by the incorporation of the CpTI gene. Intriguingly, the height and basal diameter of the clone TG04 were much greater than that of the control. The transgenic foliage consumed by insects induced the increase of larval mortality, and decrease of larval wet weight gain, faecal output, pupal weight and egg deposition. Among them 3 transgenic clones, TG04, TG07 and TG71 received special attention for their outstanding insect resistance compared with other transgenic clones, which showed that the CpTI gene in them was expressed more actively and stably than in others.展开更多
The putative transgenic hybrid triploid poplars [(P. tomentosa P. bolleana) P. tomentosa] with CpTI gene have been outplanted in test field for 2 years. Although the authors previous studies have proved that they ar...The putative transgenic hybrid triploid poplars [(P. tomentosa P. bolleana) P. tomentosa] with CpTI gene have been outplanted in test field for 2 years. Although the authors previous studies have proved that they are highly resistant to 3 species of poplar-threatening insect pests and contain high content of CpTI protein in foliage, incorporation status of foreign CpTI gene in poplar genome is uncertain. In this present study, the incorporation of foreign CpTI gene in genome of 5 transgenic poplars was confirmed by PCR and Southern blotting analysis. DNA amplification showed that there were clear DNA bands of about 450bp specific to CpTI gene in transgenic lanes, while no corresponding band in non-transgenic lane was observed. Correspondingly, clear DNA hybridization signals and no signal were exhibited on film for DNA Southern blotting analysis in transgenic lanes and non-transgenic lane, respectively, which further confirmed the stable integration of foreign CpTI gene in genome of 2-year-old transgenic poplar.展开更多
文摘In this study,cowpea trypsin inhibitor (CpTI) gene, an insecticidal gene,was introduced into two Populus tomentosa clones by gene transformation mediated by Agrobacterium tumefaciens. The transformed regeneration shoots were obtained directly by leaf discs.In order to established selection condition,leaf discs were cultured in the medium with increasing concentration kanamycin.Kanamycin resistant(km r)plantlets were obtained by 3～4 cycles screening in selective condition.Then transformed shoots were rooted in the medium containing kanamycin 50?mg/L and transferred to greenhouse.The presence of CpTI gene in transgenic plants were confirmed by PCR and PCR\|Southern blot.
文摘Both non-transgenic hybrid triploid poplars [ (Populus tomentosa×P.bolleana)×P.tomentosa ] and transgenic ones expressing cowpea trypsin inhibitor were cut at the base of the stem to produce auxoblasts, and used as source of leaves for insect feeding trials performed on 3 major insect species of poplar, the forest tent caterpillar ( Malacosoma disstria L.), gypsy moth ( Lymantria dispar L.) and willow moth ( Stilpnotia candida Staudinger). The height and basal diameter of trees were measured by the end of that year (2000). The results indicated that the growth elements of transgenic poplars were not interfered by the incorporation of the CpTI gene. Intriguingly, the height and basal diameter of the clone TG04 were much greater than that of the control. The transgenic foliage consumed by insects induced the increase of larval mortality, and decrease of larval wet weight gain, faecal output, pupal weight and egg deposition. Among them 3 transgenic clones, TG04, TG07 and TG71 received special attention for their outstanding insect resistance compared with other transgenic clones, which showed that the CpTI gene in them was expressed more actively and stably than in others.
基金Supported by the National Project in Transgenic Plant and Application (Grant No. J2002-B003)
文摘The putative transgenic hybrid triploid poplars [(P. tomentosa P. bolleana) P. tomentosa] with CpTI gene have been outplanted in test field for 2 years. Although the authors previous studies have proved that they are highly resistant to 3 species of poplar-threatening insect pests and contain high content of CpTI protein in foliage, incorporation status of foreign CpTI gene in poplar genome is uncertain. In this present study, the incorporation of foreign CpTI gene in genome of 5 transgenic poplars was confirmed by PCR and Southern blotting analysis. DNA amplification showed that there were clear DNA bands of about 450bp specific to CpTI gene in transgenic lanes, while no corresponding band in non-transgenic lane was observed. Correspondingly, clear DNA hybridization signals and no signal were exhibited on film for DNA Southern blotting analysis in transgenic lanes and non-transgenic lane, respectively, which further confirmed the stable integration of foreign CpTI gene in genome of 2-year-old transgenic poplar.
