Transformation of cucumber ( Cucumis sativus L.) plants using Agrobacterium tumefaciens and regeneration from hypocotyl explants

Nishibayashi, Soryu; Kaneko, Hiromitsu; Hayakawa, Tetsuo

doi:10.1007/s002990050126

Cited by 12 publications

(18 citation statements)

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“…Agrobacterium is presumably attracted to a wounded plant in response to signal molecules released by the plant cells to which it then becomes attracted [33,34] . The results of the present study are in agreement with previous observations [12,13,27,35] .…”

Section: Sensitivity Of Cotyledon Explants To Pptsupporting

confidence: 83%

“…Our results were in conformity with previous studies [9,10,16,26] in cucumber that 1.0 OD was essential for effective transformation. Different co-cultivation period was found optimal for the effective infection and regeneration of transgenic shoots [10,12,14,26] . The cocultivation period beyond 4 days the calluses with free of Agrobacterium contamination were more difficult to obtain for cucumber [13] .…”

Section: Sensitivity Of Cotyledon Explants To Pptmentioning

confidence: 99%

“…So far, a few reports are available on gene transfer studies of cucumber [9][10][11][12][13][14][15][16] . Based on these reports, the present study was focused on Agrobacterium-mediated transformation via direct regeneration using cotyledon explants in an important cultivar, Poinsett 76 with an important Agrobacterium tumefaciens strain EHA 105 [17].…”

Section: Introductionmentioning

confidence: 99%

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Agrobacterium-mediated Genetic Transformation in Cucumber (Cucumis sativus L.)

Ayyappan¹,

Selvaraj²,

Ganapathi³

et al. 2007

American J. of Biochemistry and Biotechnology

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The present study was undertaken to develop efficient transformation protocol for cucumber cv. Poinsett 76 using Agrobacterium strain EHA 105. Five-day-old mature cotyledon explants was used for transformation study. The infected explants were co-cultivated for 2 days in MS medium containing BA (1.0 mg L¯1). The selection of transformed shoots was carried out in MS medium fortified with BA (1.0 mg L¯1), Cefotaxime (300 mg L¯1) and PPT (2.0 mg L¯1). The transformed shoots were elongated in MS medium containing BA (1.0 mg L¯1), Cefotaxime (300 mg L¯1), PPT (2.0 mg L¯1) along with GA 3 (0.5 mg L¯1). The rooting of elongated shoots was achieved in MS medium with BA (1.0 mg L¯1), Cefotaxime (300 mg L¯1), PPT (2.0 mg L¯1) and IBA (0.6mg L¯1). The transient GUS expression assay and leaf disc assay were carried out in order to find transformed shoots. The molecular confirmation of transformed shoots revealed the foreign gene integration into cucumber genome.

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Section: Sensitivity Of Cotyledon Explants To Pptsupporting

confidence: 83%

Section: Sensitivity Of Cotyledon Explants To Pptmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Agrobacterium-mediated Genetic Transformation in Cucumber (Cucumis sativus L.)

Ayyappan¹,

Selvaraj²,

Ganapathi³

et al. 2007

American J. of Biochemistry and Biotechnology

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“…The finding of this study is in agreement with other reports for watermelon transformation (Choi et al, 1994;Park et al 2005). In many other plants, such as wheat, cucumber and Sesbania drummondii, addition of acetosyringone to the A. tumefaciens culture and/or co-cultivation medium proven to be beneficial in improving transformation efficiency (Wu et al, 2003;Nishibayashi et al, 1996;Padmanabhan and Sahi, 2009). In Astragalus racemosus, addition of acetosyringone in transformation medium showed only a small positive effect on transformation efficiency (Darlington et al, 2009).…”

Section: Effect Of Inclusion Of Acetosyringone During Co-cultivationsupporting

confidence: 82%

Cotyledon with Hypocotyl Segment as an Explant for the Production of Transgenic Citrullus vulgaris Schrad (Watermelon) Mediated by Agrobacterium tumefaciens

Suratman¹,

Huyop²,

Wagiran³

et al. 2010

Biotechnology

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Abstract:In this study, we describe the successful production of transgenic watermelon using Agrobacterium tumefaciens after optimizing a couple of transformation parameters using cotyledon with hypocotyl segment from 5 day old in vitro seedlings as target tissues. It was found that the best condition for transformation of watermelon was using bacteria at a concentration of OD 0.6, inoculation for 30 min and 600 3 day of co-cultivation. Addition of 200 µM acetosyringone in the medium helped to further increase the transformation efficiency. Agrobacterium tumefaciens strain EHA101 was found to be more effective to infect this watermelon variety as compared to LBA 4404 strain. Wounding of the explant meristems to induce multiple shoot was best carried out after 3 day. Transformed explants were selected on medium containing 5 mg LG 1 hygromycin. A total of 31 putative transformed plants were recovered with 25 shoots were found to be GUS positive after transformation of 110 cotyledons. A total of 15 GUS positive shoots amplified the hpt gene using PCR. However, only 7 shoots were successfully regenerated into whole plants.

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“…Hence, we supplemented 150 mg dm -3 kanamycin to MS medium for the selection of putative transformed explants in the subsequent experiments. Our result was justified by the previous studies where kanamycin, in the concentration of 50 -150 mg dm -3 , applied for 4 -6 weeks, was used as an efficient selection agent for cucumber transformation (Chee 1990, Nishibayashi et al 1996, Ganapathi and Perl-Treves 2000. The bacterial overgrowth during the selection of transformed explants was controlled with 300 mg dm -3 cephotaxime fortified MS medium.…”

Section: Controlmentioning

confidence: 64%

Marker-free transgenic cucumber expressing Arabidopsis cbf1 gene confers chilling stress tolerance

Gupta¹,

Rathore²,

Goyary³

et al. 2012

Biol. plant.

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Marker-free transgenic cucumber (Cucumis sativus L.) cv. Poinsett 76 SR plants were produced by Agrobacterium mediated transformation. A transformation efficiency of 1.62 was observed on using Agrobacterium tumefaciens strain LBA4404 harbouring Arabidopsis cbf1 gene driven by the inducible promoter RD29A in a binary vector system pCAMBIA. Transgene integration and single copy insert in transgenic cucumber was confirmed by polymerase chain reaction (PCR) and Southern blot analysis in T 0 lines and also confirmed marker-free status in T 1 generation. Transgene expression was confirmed by reverse transcription (RT)-PCR in T 1 generation transgenic cucumber and advanced to T 2 generation. Upon exposure to chilling stress (4 °C), the T 2 generation transgenic plants survived up to 36 h; however, wild-type plants could not survive and gradually died. A significant decrease in membrane injury index (MII), increase in activities of antioxidant enzymes (SOD and CAT), free proline content and relative water content (RWC) in the leaves were observed in transgenic cucumber as compared to wild-type under chilling stress. Thus, the transgenic cucumber plants expressing Arabidopsis cbf1 gene conferred protection against chilling stress.Additional key words: Agrobacterium, c-repeat binding factor-1, Cucumis sativus.

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Transformation of cucumber ( Cucumis sativus L.) plants using Agrobacterium tumefaciens and regeneration from hypocotyl explants

Cited by 12 publications

References 0 publications

Agrobacterium-mediated Genetic Transformation in Cucumber (Cucumis sativus L.)

Agrobacterium-mediated Genetic Transformation in Cucumber (Cucumis sativus L.)

Cotyledon with Hypocotyl Segment as an Explant for the Production of Transgenic Citrullus vulgaris Schrad (Watermelon) Mediated by Agrobacterium tumefaciens

Marker-free transgenic cucumber expressing Arabidopsis cbf1 gene confers chilling stress tolerance

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