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Strength, Stability, and cis-Motifs of In silico Identified Phloem-Specific Promoters in Brassica juncea (L.).

Koramutla MK, Bhatt D, Negi M, Venkatachalam P, Jain PK, Bhattacharya R - Front Plant Sci (2016)

Bottom Line: In RT-qPCR based gene-expression study promoter of Glutamine synthetase 3A (GS3A) showed multifold higher activity compared to others, across the different growth stages of B. juncea plants.Nevertheless, the consensus in output empirically suggested consistency in promoter-activity of the six B. juncea phloem- specific promoters including GS3A.The study identified suitable endogenous promoters for high level and consistent gene-expression in B. juncea phloem exudate.

View Article: PubMed Central - PubMed

Affiliation: National Research Centre on Plant Biotechnology, Indian Agricultural Research Institute Campus New Delhi, India.

ABSTRACT
Aphids, a hemipteran group of insects pose a serious threat to many of the major crop species including Brassica oilseeds. Transgenic strategies for developing aphid-resistant plant types necessitate phloem-bound expression of the insecticidal genes. A few known phloem-specific promoters, in spite of tissue-specific activity fail to confer high level gene-expression. Here, we identified seven orthologues of phloem-specific promoters in B. juncea (Indian mustard), and experimentally validated their strength of expression in phloem exudates. Significant cis-motifs, globally occurring in phloem-specific promoters showed variable distribution frequencies in these putative phloem-specific promoters of B. juncea. In RT-qPCR based gene-expression study promoter of Glutamine synthetase 3A (GS3A) showed multifold higher activity compared to others, across the different growth stages of B. juncea plants. A statistical method employing four softwares was devised for rapidly analysing stability of the promoter-activities across the plant developmental stages. Different statistical softwares ranked these B. juncea promoters differently in terms of their stability in promoter-activity. Nevertheless, the consensus in output empirically suggested consistency in promoter-activity of the six B. juncea phloem- specific promoters including GS3A. The study identified suitable endogenous promoters for high level and consistent gene-expression in B. juncea phloem exudate. The study also demonstrated a rapid method of assessing species-specific strength and stability in expression of the endogenous promoters.

No MeSH data available.


Related in: MedlinePlus

Significant cis-elements globally associated with phloem-specific promoters and their frequency distribution in B. juncea promoters. (A) Global occurrence of nine signature cis-elemnts in phloem-specific promoters of diverse vascular plants. (B) Frequency distribution of signature cis- elements in the seven phloem-specific promoters of B. juncea. Frequency distribution of significant motifs were searched by using FIMO program of MEME with p < 0.0001.
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Figure 5: Significant cis-elements globally associated with phloem-specific promoters and their frequency distribution in B. juncea promoters. (A) Global occurrence of nine signature cis-elemnts in phloem-specific promoters of diverse vascular plants. (B) Frequency distribution of signature cis- elements in the seven phloem-specific promoters of B. juncea. Frequency distribution of significant motifs were searched by using FIMO program of MEME with p < 0.0001.

Mentions: The global frequency of the nine identified signature cis-elements (Figure 5A) and their distribution in the seven putative phloem-specific promoters of B. juncea were analyzed (Figure 5B). The B. juncea promoters showed occurrence of all the signature cis-elements, albeit their distribution and frequency on each promoter varied (Figure 5B). Promoters of GS3A and PP2 contained eight of the nine signature motifs followed by promoter of SUC2 with six motifs. Promoters of GLP13, TGG1, and SULTR2 each contained four of the nine motifs. However, no apparent correlation between the frequency of the cis-elements and the relative strength of the promoters could be established. For example, PP2 promoter in spite of containing equal number of the motif elements as GS3A generated much lesser transcript level compared to GS3A. Interestingly, a notable difference between the GS3A and PP2 promoter was exclusive presence of a degenerate motif (GRRRGGGAGASG; R = A/G, S=G/C), identified in MEME based analysis, only in case of GS3A promoter. However, further experimentation is warranted before attributing this degenerate motif to high expression of GS3A. Nevertheless, the results of RT-qPCR analysis validated the applicability of the identified signature elements in identifying the phloem-specific promoters.


Strength, Stability, and cis-Motifs of In silico Identified Phloem-Specific Promoters in Brassica juncea (L.).

Koramutla MK, Bhatt D, Negi M, Venkatachalam P, Jain PK, Bhattacharya R - Front Plant Sci (2016)

Significant cis-elements globally associated with phloem-specific promoters and their frequency distribution in B. juncea promoters. (A) Global occurrence of nine signature cis-elemnts in phloem-specific promoters of diverse vascular plants. (B) Frequency distribution of signature cis- elements in the seven phloem-specific promoters of B. juncea. Frequency distribution of significant motifs were searched by using FIMO program of MEME with p < 0.0001.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4834444&req=5

Figure 5: Significant cis-elements globally associated with phloem-specific promoters and their frequency distribution in B. juncea promoters. (A) Global occurrence of nine signature cis-elemnts in phloem-specific promoters of diverse vascular plants. (B) Frequency distribution of signature cis- elements in the seven phloem-specific promoters of B. juncea. Frequency distribution of significant motifs were searched by using FIMO program of MEME with p < 0.0001.
Mentions: The global frequency of the nine identified signature cis-elements (Figure 5A) and their distribution in the seven putative phloem-specific promoters of B. juncea were analyzed (Figure 5B). The B. juncea promoters showed occurrence of all the signature cis-elements, albeit their distribution and frequency on each promoter varied (Figure 5B). Promoters of GS3A and PP2 contained eight of the nine signature motifs followed by promoter of SUC2 with six motifs. Promoters of GLP13, TGG1, and SULTR2 each contained four of the nine motifs. However, no apparent correlation between the frequency of the cis-elements and the relative strength of the promoters could be established. For example, PP2 promoter in spite of containing equal number of the motif elements as GS3A generated much lesser transcript level compared to GS3A. Interestingly, a notable difference between the GS3A and PP2 promoter was exclusive presence of a degenerate motif (GRRRGGGAGASG; R = A/G, S=G/C), identified in MEME based analysis, only in case of GS3A promoter. However, further experimentation is warranted before attributing this degenerate motif to high expression of GS3A. Nevertheless, the results of RT-qPCR analysis validated the applicability of the identified signature elements in identifying the phloem-specific promoters.

Bottom Line: In RT-qPCR based gene-expression study promoter of Glutamine synthetase 3A (GS3A) showed multifold higher activity compared to others, across the different growth stages of B. juncea plants.Nevertheless, the consensus in output empirically suggested consistency in promoter-activity of the six B. juncea phloem- specific promoters including GS3A.The study identified suitable endogenous promoters for high level and consistent gene-expression in B. juncea phloem exudate.

View Article: PubMed Central - PubMed

Affiliation: National Research Centre on Plant Biotechnology, Indian Agricultural Research Institute Campus New Delhi, India.

ABSTRACT
Aphids, a hemipteran group of insects pose a serious threat to many of the major crop species including Brassica oilseeds. Transgenic strategies for developing aphid-resistant plant types necessitate phloem-bound expression of the insecticidal genes. A few known phloem-specific promoters, in spite of tissue-specific activity fail to confer high level gene-expression. Here, we identified seven orthologues of phloem-specific promoters in B. juncea (Indian mustard), and experimentally validated their strength of expression in phloem exudates. Significant cis-motifs, globally occurring in phloem-specific promoters showed variable distribution frequencies in these putative phloem-specific promoters of B. juncea. In RT-qPCR based gene-expression study promoter of Glutamine synthetase 3A (GS3A) showed multifold higher activity compared to others, across the different growth stages of B. juncea plants. A statistical method employing four softwares was devised for rapidly analysing stability of the promoter-activities across the plant developmental stages. Different statistical softwares ranked these B. juncea promoters differently in terms of their stability in promoter-activity. Nevertheless, the consensus in output empirically suggested consistency in promoter-activity of the six B. juncea phloem- specific promoters including GS3A. The study identified suitable endogenous promoters for high level and consistent gene-expression in B. juncea phloem exudate. The study also demonstrated a rapid method of assessing species-specific strength and stability in expression of the endogenous promoters.

No MeSH data available.


Related in: MedlinePlus