Dataset Information

Structural and Functional Analysis of a Bidirectional Promoter from Gossypium hirsutum in Arabidopsis.

ABSTRACT: Stacked traits have become an important trend in the current development of genomically modified crops. The bidirectional promoter can not only prevent the co-suppression of multigene expression, but also increase the efficiency of the cultivation of transgenic plants with multigenes. In Gossypium hirsutum, Ghrack1 and Ghuhrf1 are head-to-head gene pairs located on chromosome D09. We cloned the 1429-bp intergenic region between the Ghrack1 and Ghuhrf1 genes from Gossypium hirsutum. The cloned DNA fragment GhZU had the characteristics of a bidirectional promoter, with 38.7% G+C content, three CpG islands and no TATA-box. Using gfp and gus as reporter genes, a series of expression vectors were constructed into young leaves of tobacco. The histochemical GUS (Beta-glucuronidase) assay and GFP (green fluorescence protein) detection results indicated that GhZU could drive the expression of the reporter genes gus and gfp simultaneously in both orientations. Furthermore, we transformed the expression vectors into Arabidopsis and found that GUS was concentrated at vigorous growth sites, such as the leaf tip, the base of the leaves and pod, and the stigma. GFP was also mainly expressed in the epidermis of young leaves. In summary, we determined that the intergenic region GhZU was an orientation-dependent bidirectional promoter, and this is the first report on the bidirectional promoter from Gossypium hirsutum. Our findings in this study are likely to enhance understanding on the regulatory mechanisms of plant bidirectional promoters.

SUBMITTER: Yang J

PROVIDER: S-EPMC6274729 | biostudies-literature | 2018 Oct

REPOSITORIES: biostudies-literature

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Structural and Functional Analysis of a Bidirectional Promoter from Gossypium hirsutum in Arabidopsis.

Yang Jiangtao J Wang Xujing X Hasi Agula A Wang Zhixing Z

International journal of molecular sciences 20181023 11

Stacked traits have become an important trend in the current development of genomically modified crops. The bidirectional promoter can not only prevent the co-suppression of multigene expression, but also increase the efficiency of the cultivation of transgenic plants with multigenes. In Gossypium hirsutum, Ghrack1 and Ghuhrf1 are head-to-head gene pairs located on chromosome D09. We cloned the 1429-bp intergenic region between the Ghrack1 and Ghuhrf1 genes fro ...[more]

PMID: 30360512

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Project description:Apoplastic ascorbate oxidase (AO) plays significant roles in plant cell growth. However, the mechanism of underlying the transcriptional regulation of AO in Gossypium hirsutum remains unclear. Here, we obtained a 1,920-bp promoter sequence from the Gossypium hirsutum ascorbate oxidase (GhAO1) gene, and this GhAO1 promoter included a number of known cis-elements. Promoter activity analysis in overexpressing pGhAO1::GFP-GUS tobacco (Nicotiana benthamiana) showed that the GhAO1 promoter exhibited high activity, driving strong reporter gene expression in tobacco trichomes, leaves and roots. Promoter 5'-deletion analysis demonstrated that truncated GhAO1 promoters with serial 5'-end deletions had different GUS activities. A 360-bp fragment was sufficient to activate GUS expression. The P-1040 region had less GUS activity than the P-720 region, suggesting that the 320-bp region from nucleotide -720 to -1040 might include a cis-element acting as a silencer. Interestingly, an auxin-responsive cis-acting element (TGA-element) was uncovered in the promoter. To analyze the function of the TGA-element, tobacco leaves transformed with promoters with different 5' truncations were treated with indole-3-acetic acid (IAA). Tobacco leaves transformed with the promoter regions containing the TGA-element showed significantly increased GUS activity after IAA treatment, implying that the fragment spanning nucleotides -1760 to -1600 (which includes the TGA-element) might be a key component for IAA responsiveness. Analyses of the AO promoter region and AO expression pattern in Gossypium arboreum (Ga, diploid cotton with an AA genome), Gossypium raimondii (Gr, diploid cotton with a DD genome) and Gossypium hirsutum (Gh, tetraploid cotton with an AADD genome) indicated that AO promoter activation and AO transcription were detected together only in D genome/sub-genome (Gr and Gh) cotton. Taken together, these results suggest that the 1,920-bp GhAO1 promoter is a functional sequence with a potential effect on fiber cell development, mediated by TGA-element containing sequences, via the auxin-signaling pathway.

Dataset Information

Structural and Functional Analysis of a Bidirectional Promoter from Gossypium hirsutum in Arabidopsis.

Publications

Structural and Functional Analysis of a Bidirectional Promoter from <i>Gossypium hirsutum</i> in <i>Arabidopsis</i>.

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