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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.


Gene ontology of phloem-specific transcripts. (A) Nine functional categories of the transcripts are represented by different letters in the pie chart. I. Response to biotic and abiotic stimulus, II. Developmental processes, III. Transcription, IV. Transport, V. Signal transduction, VI. Electron transport, VII. Cell organization and biogenesis, VIII. Protein metabolism, and IX. Unknown biological processes. (B) Distribution of Go terms based on their molecular function.
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Figure 1: Gene ontology of phloem-specific transcripts. (A) Nine functional categories of the transcripts are represented by different letters in the pie chart. I. Response to biotic and abiotic stimulus, II. Developmental processes, III. Transcription, IV. Transport, V. Signal transduction, VI. Electron transport, VII. Cell organization and biogenesis, VIII. Protein metabolism, and IX. Unknown biological processes. (B) Distribution of Go terms based on their molecular function.

Mentions: Based on literature, 39 known phloem-specific promoters were chosen and their sequences were downloaded from NCBI database (Table S3). These promoters, from monocot as well as dicot plants, ranged from smallest AtGLP13-promoter of 762 bp to CmGAS1-promoter of 3000 bp. Based on putative function of their cognate genes, the promoters were classified in two GO terms viz. biological process (Figure 1A) and molecular function (Figure 1B). In biological process among the major categories, about 26.6% of the promoters were found to be stress responsive and grouped under response to stresses, followed by 20%, that were associated with developmental processes, and 10% each controlling genes related to transport and transcription processes. Across the ontogenic groups, seven promoters that were associated with host-response against insects and pathogens, either directly or indirectly, were identified (Table 1). Homologues of the seven promoters were predicted in Brassica spp. and identified either in B. rapa or in B. juncea.


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)

Gene ontology of phloem-specific transcripts. (A) Nine functional categories of the transcripts are represented by different letters in the pie chart. I. Response to biotic and abiotic stimulus, II. Developmental processes, III. Transcription, IV. Transport, V. Signal transduction, VI. Electron transport, VII. Cell organization and biogenesis, VIII. Protein metabolism, and IX. Unknown biological processes. (B) Distribution of Go terms based on their molecular function.
© Copyright Policy
Related In: Results  -  Collection

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

Figure 1: Gene ontology of phloem-specific transcripts. (A) Nine functional categories of the transcripts are represented by different letters in the pie chart. I. Response to biotic and abiotic stimulus, II. Developmental processes, III. Transcription, IV. Transport, V. Signal transduction, VI. Electron transport, VII. Cell organization and biogenesis, VIII. Protein metabolism, and IX. Unknown biological processes. (B) Distribution of Go terms based on their molecular function.
Mentions: Based on literature, 39 known phloem-specific promoters were chosen and their sequences were downloaded from NCBI database (Table S3). These promoters, from monocot as well as dicot plants, ranged from smallest AtGLP13-promoter of 762 bp to CmGAS1-promoter of 3000 bp. Based on putative function of their cognate genes, the promoters were classified in two GO terms viz. biological process (Figure 1A) and molecular function (Figure 1B). In biological process among the major categories, about 26.6% of the promoters were found to be stress responsive and grouped under response to stresses, followed by 20%, that were associated with developmental processes, and 10% each controlling genes related to transport and transcription processes. Across the ontogenic groups, seven promoters that were associated with host-response against insects and pathogens, either directly or indirectly, were identified (Table 1). Homologues of the seven promoters were predicted in Brassica spp. and identified either in B. rapa or in B. juncea.

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.