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United States Department of Agriculture

Agricultural Research Service

Title: Genetic Transformation of Durum Wheat: Expression of Transgenes in T0 and T1 Generations.

Authors
item Bommineni, Venkata
item Jauhar, Prem
item Peterson, Terrance - NDSU, PLNT SCI., FARGO,ND

Submitted to: Plant Genome Conference Proceedings
Publication Type: Proceedings
Publication Acceptance Date: December 20, 1995
Publication Date: N/A

Technical Abstract: Durum wheat (Triticum turgidum L., 2n=4x=28: AABB genomes) is widely used for making pasta and semolina, however, the usefulness of biotechnological tools in germplasm enhancement of durum have not been explored. Our objective was to develop a routine gene delivery system in some agronomically superior durum cultivars such as Medora, Monroe, Renville and Vic. First, we established an efficient method of regenerating plantlets through scutellum culture. Isolated scutella were then used as target cells to deliver gus, nptII and bar genes, using a biolistic device. A high GUS activity was observed two days after bombardment with pRC62 and pBARGUS DNA vectors. The bombarded scutella were cultured on a callus induction medium for up to six weeks and then transferred to an appropriate selection medium and incubated for 4-6 weeks. Geneticin-418 at 70 mg/L was used to recover the putative transformants incorporating nptII, whereas L-ppt (Basta) at 5 mg/L was used to select the transformants containing the bar gene. The resistant plantlets thus recovered were transferred to non-selective medium for 2-3 weeks to establish before planting them into greenhouse pots. Following this procedure, a transformation frequency of 0.9% with pRC62 (cv. Monroe) and 1.2% with pBARGUS (cv. Medora) was obtained. Expression of gus was monitored histochemically in leaves, flower organs and brush portions of mature seeds to T0 and T1 plants. The incorporation of bar was verified by spot brushing the leaves of the plants derived from L-ppt selection medium or by spraying the whole T1 plants with Basta solution (120 mg/L of L-ppt). The integration of gus and bar genes into the genomes of durum wheat cultivars was confirmed by Southern blot analyses.

Last Modified: 7/30/2014
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