Establishment of apple hairy root culture by Agrobacterium rhizogenes vector carrying the serine proteinase inhibitor gene
2013
Аутори:
Stanišić, MarianaBanjac, Nevena
Savić, Jelena
Ćosić, Tatjana
Smigocki, Ann
Ninković, Slavica
Остала ауторства
Vinterhalter, DraganТип документа:
Конференцијски прилог (Објављена верзија)
,
© 2013 by the Serbian Plant Physiology Society
Метаподаци
Приказ свих података о документуАпстракт:
Melrose is the economically important apple (Malus domestica Borkh.) cultivar grown worldwide. Susceptibility to insect pests of Melrose and other apple cultivars could be reduced by genetic transformation with genes coding proteins with insecticidal properties. The use of proteinase inhibitor genes could be the promising alternative to enhance tolerance to insect pests. In vitro-grown young shoots of apple cultivar Melrose were transformed using Agrobacterium rhizogenes agropine strain 15834 containing the pCAMBIA 1301 binary vector carrying the serine proteinase inhibitor gene from Beta vulgaris L. The shoots were inoculated by wounding with a sterile needle immersed in overnight bacterial suspension. After three days of co-cultivation on hormone-free MS medium, shoots were transferred onto hormone-free MS medium supplemented with 500 mg/l cefotaxime. Whitish and thick transformed roots appeared from wounds three to four weeks after the inoculation. The efficiency of transformation was 22.2%, calculated as a number of shoots producing at least one transgenic root. The PCR analysis indicated variability in the integration of rolA,B,C,D genes in the genomic DNA of transformed roots. The rolC and the rolD genes were present in all, but the rolA gene was presented in six, and the rolB in five of seven hairy root clones. Transformed roots had no the ags or the aux1 genes. Excised roots grew only on liquid MS medium supplemented with 0.05 mg/1 IBA. We assume that it is the result of roots inability to produce adequate levels of endogenous auxins because of the lack of the aux1 gene. The RT-PCR analysis showed expression of proteinase inhibitor gene in two of seven transformed clones. This protocol is the bases for future transformation experiments with other apple cultivars and different proteinase inhibitor genes under control of constitutive and inducible promoters.
Финансирање / пројекти:
- Биотехнологија ин витро - гајене, лековите и угрожене биљне врсте (RS-MESTD-Basic Research (BR or ON)-173015)
У:
- Vinterhalter D, editor. Programme and Abstracts: 1st International Conference on Plant Biology and 20th Symposium of the Serbian Plant Physiology Society; 2013 Jun 4-7; Subotica, Belgrade. Belgrade: Serbian Plant Physiology Society; 2013. p. 65-6.