Tobacco transformation and analysis

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Tobacco transformation and analysis

Written by nguyen

Tuesday, 27 January 2009 05:00

Various stages of transformation and analysis of transgene expression.

A. Leaf dishes after co-cultured (top panel) and regeneration of plants (bottom panel-left) from leaf dishes on selection medium (hygromycin 30 mg/l); Petri at right bottom corner contain wild type that no co-cultured with Agrobacterium.

B. Responding of shoots (top panel) and roots (bottom panel) of transgenic plants (left) and wide type plant (right) on selection medium.
C. Transgenic tobacco plants in pots.
D. Transgenic plant in flowering stage in the greenhouse.
E. Germination of transgenic (top panel) and wild type (bottom) seeds on selection medium (hygromycin 30 mg/l).
F. Confirmation GUS gene in seeds (top) and seedling (bottom), brown seed are wild type.

Difference responding of transgenic tobacco plants those transformed with different constructs.

A. Plants those transformed with pITB250, pITB228 and pBin-Tet/pITB228, respectively (left to right)
B. Plants those transformed with pITB550, pITB650 (top), pITB541 and pITB641 (bottom).
C. Plants those transformed with pITB750, pITB850 (top), pITB741 and pITB841 (bottom).
D. Plants those transformed with pITB450 and pITB441 (left to right).
E. Plants those transformed with pCAMBIA1301 (left) and pBin-Tet/pITB228 (right).
F. Flowering of plants those transformed with pCAMBIA1301 (left), pITB228 (middle) and pBin-Tet/pITB228 (right).

Histochemical localization of GUS expression in transgenic tobacco callus (A) and shoot (B) which were transformed with construct pITB441 (Nt.441) and in shoots transformed with construct pITB450 (C). The leaf transformed with pITB641 (D) or pITB650 (E) showing more GUS expression in guard cells. While leaf transformed with construct pITB750 (F) shown highest expression in vascular and lower in mesophyll tissues, in contrast leaf transformed with construct pITB450 (G) showing more expression in mesophyll and absence in vascular tissues. In the roots, expression of GUS gene is specific; the root transformed with construct pITB741 (H) or pITB750 (I) is high in vascular tissues, while with pITB541 (J) or pITB550 (K) were not expression in the root hair (RH) and root elongation (RE) but more in root cap (RC); with pITB228 (L) GUS expression was in whole root.