Figure 1.
Gene construct: pKanIntron-35S-SHI used for Agrobacterium-mediated transformation of poinsettia.
Table 1.
Primers for real-time PCR expression analysis of AtSHI in poinsettia.
Figure 2.
PCR analysis of poinsettia transformed with the AtSHI gene (genomic DNA was extracted from leaves).
Lane 1∶100 bp marker, lane 2: plasmid control, lane 3: blank, lanes 4–11: eight plants from independent transgenic lines 1 (TL1) (individuals 1–8), lane 12: TL2, lane 13: TL3 and lane 14: WT control line. The arrow indicates the 500 bp band.
Figure 3.
Southern blot analysis of PCR positive transgenic poinsettia lines overexpressing the AtSHI gene.
HindIII-digested total genomic DNAs were hybridized with a AtSHI probe (the 500 bp PCR product). Lane 1: positive control, the 500 bp PCR product; lane 2–4: TL1–3, TL1–4, TL1–6 of the transgenic line one (TL1); lane 5: TL2; lane 6: TL3, lane 7: WT control.
Figure 4.
Quantitative real-time PCR analysis of AtSHI transgene in different transgenic lines of poinsettia.
Two microgram total RNAs from transgenic poinsettia lines and the endogenous control, α-tubulin gene were used for synthesize cDNAs prior the real-time qPCR analysis. Values are means of three technical replications except TL2 (n = 2). Data were analyzed using the 2−△△CT method and represented as log10 values. Vertical bars represent the ± SE (standard error).
Figure 5.
Height comparison among the different transgenic lines (TL) of AtSHI overexpressing poinsettia and untransformed control plants grown under short day (10 h) (A) and long day conditions (16 h) (B).
Vertical bars represent the ± SE (standard error), n = 11–12 and 6–12 in A and B, respectively.
Figure 6.
Transgenic AtSHI overexpressing poinsettia plants (A), petioles (B) and leaves (C).
In each figure: from the left different transgenic lines; TL1, TL2, TL3 and non-transformed control plants are shown. The plants were grown at 21±2°C, a 10 h photoperiod at an irradiance of 100±20 µmol m−2 s−1.
Figure 7.
Effects of AtSHI overexpression on internode length (A), total internode number (sum of bracts and leaves) (B), bract number (C) and bract area (D) of different transgenic lines and control plants of poinsettia.
Plants were grown under short day conditions of a 10 h photoperiod, n = 11–12. Mean values with different letters are significantly different based on ANOVA followed by a Tukey’s test at p≤0.05. Vertical bars represent the ± SE (standard error).
Table 2.
Comparison of growth parameters among transgenic (T) lines of poinsettia overexpressing the AtSHI gene, TL1, TL2, TL3 and control plants under short day conditions of a 10 h photoperiod in a controlled environment.
Table 3.
Endogenous levels of auxin and their metabolites (pmol g−1 DW) in transgenic lines and control plants of poinsettia grown under a short photoperiod of 10 h.