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Identification and functional analysis of pistil self-incompatibility factor HT-B of Petunia.

Puerta AR, Ushijima K, Koba T, Sassa H - J. Exp. Bot. (2009)

Bottom Line: The structural features and expression pattern of Petunia HT-B were very similar to those of Nicotiana and Solanum.Unlike other solanaceous species, expression of HT-B was also observed in self-compatible Petunia species.Quantitative analysis of the HT-B mRNA accumulation in the transgenics showed that a 100-fold reduction is not sufficient and a >1000-fold reduction is required to achieve partial breakdown of GSI.

View Article: PubMed Central - PubMed

Affiliation: Graduate School of Science and Technology, Chiba University, 648 Matsudo, Matsudo, Chiba 271-8510, Japan.

ABSTRACT
Gametophytic self-incompatibility (GSI) in Solanaceae, Rosaceae, and Plantaginaceae is controlled by a multiallelic S-locus. The specificities of pistil and pollen are controlled by separate S-locus genes, S-RNase and SLF/SFB, respectively. Although the S-specificity is determined by the S-locus genes, factors located outside the S-locus are also required for expression of GSI. HT-B is one of the pistil non-S-factors identified in Nicotiana and Solanum, and encodes a small asparagine/aspartate-rich extracellular protein with unknown biochemical function. Here, HT-B was cloned from Petunia and characterized. The structural features and expression pattern of Petunia HT-B were very similar to those of Nicotiana and Solanum. Unlike other solanaceous species, expression of HT-B was also observed in self-compatible Petunia species. RNA interference (RNAi)-mediated suppression of Petunia HT-B resulted in partial breakdown of GSI. Quantitative analysis of the HT-B mRNA accumulation in the transgenics showed that a 100-fold reduction is not sufficient and a >1000-fold reduction is required to achieve partial breakdown of GSI.

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Related in: MedlinePlus

RNA blot analysis of S3L-RNase in styles of RNAi lines for HT-B. Severely HT-B-suppressed Mitchell background plants 52 and 208 were analysed. ‘Mitchell’ and untransformed Mitchell background (CM) are negative and positive controls, respectively. A 10 μg aliquot of style total RNA was loaded in each lane, blotted, and hybridized with the P. inflata S3L-RNase probe. The ethidium bromide-stained gel is shown to ascertain equal loading conditions.
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fig9: RNA blot analysis of S3L-RNase in styles of RNAi lines for HT-B. Severely HT-B-suppressed Mitchell background plants 52 and 208 were analysed. ‘Mitchell’ and untransformed Mitchell background (CM) are negative and positive controls, respectively. A 10 μg aliquot of style total RNA was loaded in each lane, blotted, and hybridized with the P. inflata S3L-RNase probe. The ethidium bromide-stained gel is shown to ascertain equal loading conditions.

Mentions: Real-time PCR was performed in selected plants to quantitate the level of reduction of HT-B gene expression. As shown in Fig. 8A, PiHT-B expression in the partial SC ‘Mitchell’ background plants 52 and 208 was reduced by >1000 times in comparison with the control (CM). Suppression in hybrid background plants 78, 87, and 176 was >100 times. The double transformant (plant 94) presented a suppression of >100 times for both genes (Fig. 8A, B). Unaffected expression of S3L-RNase in the partial SC plants 52 and 208 was confirmed through RNA blot analysis including an untransformed plant (CM) and ‘Mitchell’ as positive and negative controls, respectively (Fig. 9).


Identification and functional analysis of pistil self-incompatibility factor HT-B of Petunia.

Puerta AR, Ushijima K, Koba T, Sassa H - J. Exp. Bot. (2009)

RNA blot analysis of S3L-RNase in styles of RNAi lines for HT-B. Severely HT-B-suppressed Mitchell background plants 52 and 208 were analysed. ‘Mitchell’ and untransformed Mitchell background (CM) are negative and positive controls, respectively. A 10 μg aliquot of style total RNA was loaded in each lane, blotted, and hybridized with the P. inflata S3L-RNase probe. The ethidium bromide-stained gel is shown to ascertain equal loading conditions.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC2657546&req=5

fig9: RNA blot analysis of S3L-RNase in styles of RNAi lines for HT-B. Severely HT-B-suppressed Mitchell background plants 52 and 208 were analysed. ‘Mitchell’ and untransformed Mitchell background (CM) are negative and positive controls, respectively. A 10 μg aliquot of style total RNA was loaded in each lane, blotted, and hybridized with the P. inflata S3L-RNase probe. The ethidium bromide-stained gel is shown to ascertain equal loading conditions.
Mentions: Real-time PCR was performed in selected plants to quantitate the level of reduction of HT-B gene expression. As shown in Fig. 8A, PiHT-B expression in the partial SC ‘Mitchell’ background plants 52 and 208 was reduced by >1000 times in comparison with the control (CM). Suppression in hybrid background plants 78, 87, and 176 was >100 times. The double transformant (plant 94) presented a suppression of >100 times for both genes (Fig. 8A, B). Unaffected expression of S3L-RNase in the partial SC plants 52 and 208 was confirmed through RNA blot analysis including an untransformed plant (CM) and ‘Mitchell’ as positive and negative controls, respectively (Fig. 9).

Bottom Line: The structural features and expression pattern of Petunia HT-B were very similar to those of Nicotiana and Solanum.Unlike other solanaceous species, expression of HT-B was also observed in self-compatible Petunia species.Quantitative analysis of the HT-B mRNA accumulation in the transgenics showed that a 100-fold reduction is not sufficient and a >1000-fold reduction is required to achieve partial breakdown of GSI.

View Article: PubMed Central - PubMed

Affiliation: Graduate School of Science and Technology, Chiba University, 648 Matsudo, Matsudo, Chiba 271-8510, Japan.

ABSTRACT
Gametophytic self-incompatibility (GSI) in Solanaceae, Rosaceae, and Plantaginaceae is controlled by a multiallelic S-locus. The specificities of pistil and pollen are controlled by separate S-locus genes, S-RNase and SLF/SFB, respectively. Although the S-specificity is determined by the S-locus genes, factors located outside the S-locus are also required for expression of GSI. HT-B is one of the pistil non-S-factors identified in Nicotiana and Solanum, and encodes a small asparagine/aspartate-rich extracellular protein with unknown biochemical function. Here, HT-B was cloned from Petunia and characterized. The structural features and expression pattern of Petunia HT-B were very similar to those of Nicotiana and Solanum. Unlike other solanaceous species, expression of HT-B was also observed in self-compatible Petunia species. RNA interference (RNAi)-mediated suppression of Petunia HT-B resulted in partial breakdown of GSI. Quantitative analysis of the HT-B mRNA accumulation in the transgenics showed that a 100-fold reduction is not sufficient and a >1000-fold reduction is required to achieve partial breakdown of GSI.

Show MeSH
Related in: MedlinePlus