Transient expression assay for the optimization of direct gene transfer into cucumber meristem protoplasts by electroporation

• Authors: W. Burza , P. Wochniak , T. Wroblewski , S. Malepszy
• Languages of publication: EN

Abstracts

EN

The paper obtained a new way of viable and very homogenous cucumber protoplasts.Protoplasts from cells formed in the shoot tip meristem culture were isolated from suspension.Plasmid pBI121 was introduced using imulse electric field.Effectiveness of transformation process was determinated by transient expression of ?-glucoronidase (GUS) gene, controlled by promotor 35S.The ativity of ?-glucoronidase enzyme as a product of GUS reporter gene was estimated by fluorometric method (Jefferson 1987).Parameters of electroporatian process were optimized.The transient expression of GUS gene was measured 24 h after electroporation.The highest effectiveness of transformation process was achieved using three electric impulses at the initial voltage of 250-350V at 10-sec.intervals as a results of discharging a 140 ?F capacitor and 50-70 ?g x cm/1000 plasmid DNA in the presence of 50 ?g x cm/1000 carrier DNA.The system presented is an effective method of exogenic DNA transfer, which is indicated by a high transient expression of the reporter gene.In comparison to Agrobacterium tumefaciens and A.rhizogenes, this alternative method of gene transfer can be used for obtaining transgenic cucumber plants.

Keywords

EN

CUCUMIS SATIVUS; ELECTROPORATION; TRANSIENT EXPRESSION

Discipline

• GENETICS: GENETICS

• Publisher: Institute of Plant Genetics, Polish Academy of Sciences
• Journal: Journal of Applied Genetics
• Year: 1995
• Volume: 36
• Issue: 1
• Pages: 1-10

Contributors

author

W. Burza

author

P. Wochniak

author

T. Wroblewski

author

S. Malepszy

• Document Type: article
• Publication order reference: W.Burza