Fig 1.
Detection of StGCS-GS transcripts by semi-quantitative RT-PCR in transgenic sugar beets.
(a) Semi-quantitative RT-PCR analysis of seven transgenic sugar beet (Beta vulgaris L.) lines overexpressing StGCS-GS (s1 to s7). BvActin1 was used as a loading control. The PCR products were separated by electrophoresis on agarose gels and visualized by staining with ethidium bromide. (b) The intensity of each band was measured using Quantity One software. The blank bars represent the StGCS-GS/BvActin1 expression intensity ratios. Different italicized letters (a-c) indicate expression differences (P<0.05) between lines. wt: wild-type.
Fig 2.
Enhancement of heavy metal tolerance by overexpression of StGCS-GS in transgenic sugar beets.
One-week-old transgenic seedlings (s2, s4 and s5) and wild-type seedlings (wt) were grown for 3 weeks on MS medium +1.5 mg L-1 NAA supplemented with 0 (Control), 50, 100 or 200 μM CdCl2, ZnCl2 or CuCl2.
Fig 3.
Comparative analysis of the root length and fresh weight of transgenic lines under heavy metal stress.
Root lengths (a, c and e) and fresh weights (b, d and f) of transgenic sugar beets overexpressing StGCS-GS (s2, s4 and s5) and wild-type sugar beets (wt) were measured after 3 weeks of treatment with 0, 50, 100 or 200 μM Cd2+, Zn2+ or Cu2+. The experiments were performed in triplicate, and the values are presented as the means ± SD. Means denoted by the same italicized letter did not differ significantly (P<0.05).
Fig 4.
Growth inhibition of transgenic lines by heavy metal ions under hydroponic conditions.
Both transgenic (s1, s2 and s3) and wild-type (wt) sugar beets were cultivated in one-half-strength Hoagland solution for 3 weeks and treated with 0, 50, 100 or 200 μM Cd (a), Zn (b) and Cu (c) for 14 days. Relative growth was expressed as the percentage increase in fresh weight of the same line during this period. The values shown are the average ± standard deviation. Significant differences (P<0.05) are indicated as italicized letters (a-d).
Fig 5.
Heavy metal ion accumulation of transgenic sugar beets under Cd, Zn and Cu stress.
The cadmium, zinc and copper content of shoots (a, c and e) and roots (b, d and f) of different transgenic lines was determined after exposure to 50, 100 or 200 μM CdCl2, ZnCl2 or CuCl2 for 3 weeks. wt: wild-type; s2, s4 and s5: different transgenic lines overexpressing StGCS-GS. The values shown are the means ± standard deviation. Different italicized letters (a-d) indicate significant differences (P<0.05) between lines.
Fig 6.
Glutathione and phytochelator content in different transgenic lines exposed to cadmium, zinc and copper.
The total glutathione (GSH) (a) and phytochelator (PC) (b) contents of the transgenic lines were determined before (control) and after exposure to 100 μM CdCl2, ZnCl2 or CuCl2 for 3 weeks. wt: wild-type; s2, s4 and s5: transgenic lines overexpressing StGCS-GS. The values shown are means ± standard deviation. Different italicized letters (a-c) at the top of the error bars indicate significant differences (P<0.05).
Fig 7.
Multiple heavy metal tolerance of transgenic sugar beets overexpressing StGCS-GS.
(a) One-week-old transgenic sugar beet lines overexpressing StGCS-GS (s2, s4 and s5) were grown on rooting medium (MS + 1.5 mg L-1 NAA) supplemented with combinations of two or three ions of 50 μM Cd, Zn or Cu (Cd-Zn, Cd-Cu, Zn-Cu or Cd-Zn-Cu) for 3 weeks. The wild-type seedlings (wt) were used as controls. (b) The fresh weight of all tested lines was measured three times, and the results are presented as the means ± SD. Different italicized letters indicate significant differences (P<0.05).
Table 1.
Accumulation of Cd, Zn and Cu ions in transgenic sugar beet shoots under complex heavy metal stress.
Table 2.
Accumulation of Cd, Zn and Cu ions in the roots of transgenic sugar beets under complex heavy metal stress.