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Acta Horticulturae
Pepper (Capsicum annuum L.) is widely considered as a recalcitrant species for genetic transformation studies. Plant regeneration via adventitious organogenesis in pepper, which usually occurs at low frequencies, is genotype-dependent. Interestingly, a SEM study that we conducted on cotyledon explants shows that bud induction occurs at a high incidence. The buds seem to arise directly from subepidermal cell layers near the wound surface, but not from callus tissue that may develop on the explant. Commonly, buds develop into adventitious leaf structures ("blind leaves") rather than into normal shoots. We did not find in our SEM study typical domes of apical shoot meristems. Thus, the scarce plant regeneration seems to be a consequence of sporadic differentiation of normal apical shoot meristems, but not due to a defect in shoot development from available meristems. We assessed the efficiency of Agrobacterium - mediated transformation by examining transient expression of GUS or GFP. Transient cell transformation was observed at high frequencies, particularly following ultrasonication of cotyledons. Stable expression was obtained in a much smaller fraction of the transformed cells. The use of GFP enabled to reveal many transgenic calli along wounds or on the surface of explants. However, we normally couldn't obtain transformed adventitious leaf structures. These results indicate that no spatial overlap exists between regeneration-competent cells and those cells that undergo stable Agrobacterium - mediated transformation.
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Why is it difficult to obtain transgenic pepper plants?
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Why is it difficult to obtain transgenic pepper plants?
Pepper (Capsicum annuum L.) is widely considered as a recalcitrant species for genetic transformation studies. Plant regeneration via adventitious organogenesis in pepper, which usually occurs at low frequencies, is genotype-dependent. Interestingly, a SEM study that we conducted on cotyledon explants shows that bud induction occurs at a high incidence. The buds seem to arise directly from subepidermal cell layers near the wound surface, but not from callus tissue that may develop on the explant. Commonly, buds develop into adventitious leaf structures ("blind leaves") rather than into normal shoots. We did not find in our SEM study typical domes of apical shoot meristems. Thus, the scarce plant regeneration seems to be a consequence of sporadic differentiation of normal apical shoot meristems, but not due to a defect in shoot development from available meristems. We assessed the efficiency of Agrobacterium - mediated transformation by examining transient expression of GUS or GFP. Transient cell transformation was observed at high frequencies, particularly following ultrasonication of cotyledons. Stable expression was obtained in a much smaller fraction of the transformed cells. The use of GFP enabled to reveal many transgenic calli along wounds or on the surface of explants. However, we normally couldn't obtain transformed adventitious leaf structures. These results indicate that no spatial overlap exists between regeneration-competent cells and those cells that undergo stable Agrobacterium - mediated transformation.
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