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Transgenic Date Palm Containing Endotoxin Cry3Aa Gene

Journal of Agricultural Science and Technology(A) 2017年第4期7卷 246-257页

作者： Awatef Mahmoud Badr-Elden Ibrahim Abd El-Maksoud Ibrahim Hamdy ahmed Emaral Mahdia Farid Gabr ahmed abbas nower Tamer Mahfouz Abd Elaziem Department of Plant Biotechnology Genetic Engineering and Biotechnology Research Institute (GEBRI) University of Sadat CitySadat City 32897 Egypt Department of Plant Genetic Resources Desert Research Center Cairo 11753 Egypt

Date palm, like all other crops, is very sensitive to the injury by many insect pests, which may lead to the death of the affected plant and causes decrease in yield. In the present study, an efficient Agrobacterium for genetic transformation was successfully achieved for well known date palm （Phoenix dactylifera L.） cv. Medjool and Khalas using callus as explant. Embryogenic callus were recorded 100% mortality when cultured on MS medium containing 100 mg/L kanamycin with different cultivars, thus it was chosen for the selection of transformed explants. Embryogenic callus of Medjool and Khalas were incubated with Agrobacterium tumefaciens strain LBA 4404 for 0.5, 1, 2, 4 and 24 h on LB medium. After the incubation periods, embryogenic callus was transferred to MS medium with 0.1 mg/L NAA, 0.05 mg/L BA, 250 mg/L carbenicillin and 100 mg/L kanamycin for detection of transgenic embryogenic callus. Polymerase chain reaction （PCR） was used for the rapid screening of Cry3Aa gene. For screening, total genomic DNA was isolated from transformants. Using primer specific to Cry3Aa gene （forward and reverse）, a PCR product with a size of about 2,000 bp was amplified when all nucleic acid from the transformants were utilized as templates. PCR analysis confirmed the appearance of the transgene of 2,000 bp in one individual plantlet. Presence and integration of foreign Cry3Aa gene in regenerated kanamycin resistant embryogenic callus was also confirmed by Southern blot hybridization. It was found that one transgenic embryogenic callus for both Medjool and Khalas showed a single copy of gene integration. These results signify the successful transfer of Cry3Aa gene into date palm plant.

关键词： Date palm Agrobacterium tumefaciens Cry3Aa gene Southern blot analysis transformation.

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