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New insights into plant glycoside hydrolase family 32 in Agave species.

Avila de Dios E, Gomez Vargas AD, Damián Santos ML, Simpson J - Front Plant Sci (2015)

Bottom Line: Notably however neither 1-FFT nor 6-SFT encoding cDNAs were identified.In silico analysis revealed that distinct isoforms for certain enzymes found in a single species, showed different levels and tissue specific patterns of expression whereas in other cases expression patterns were conserved both within the species and between different species.Thin layer chromatography confirmed the presence of fructans with degree of polymerization (DP) greater than DP three in both immature buds and fully opened flowers also obtained from A. tequilana.

View Article: PubMed Central - PubMed

Affiliation: Department of Genetic Engineering, Centro de Investigación y Estudios Avanzados Irapuato, Mexico.

ABSTRACT
In order to optimize the use of agaves for commercial applications, an understanding of fructan metabolism in these species at the molecular and genetic level is essential. Based on transcriptome data, this report describes the identification and molecular characterization of cDNAs and deduced amino acid sequences for genes encoding fructosyltransferases, invertases and fructan exohydrolases (FEH) (enzymes belonging to plant glycoside hydrolase family 32) from four different agave species (A. tequilana, A. deserti, A. victoriae-reginae, and A. striata). Conserved amino acid sequences and a hypervariable domain allowed classification of distinct isoforms for each enzyme type. Notably however neither 1-FFT nor 6-SFT encoding cDNAs were identified. In silico analysis revealed that distinct isoforms for certain enzymes found in a single species, showed different levels and tissue specific patterns of expression whereas in other cases expression patterns were conserved both within the species and between different species. Relatively high levels of in silico expression for specific isoforms of both invertases and fructosyltransferases were observed in floral tissues in comparison to vegetative tissues such as leaves and stems and this pattern was confirmed by Quantitative Real Time PCR using RNA obtained from floral and leaf tissue of A. tequilana. Thin layer chromatography confirmed the presence of fructans with degree of polymerization (DP) greater than DP three in both immature buds and fully opened flowers also obtained from A. tequilana.

No MeSH data available.


qRT-PCR analysis of transcripts encoding 1-SST, 6G-FFT, and Cwinv type enzymes in different floral tissues of A. tequilana. (A) Example of flower buds and floral tissues used for qRT-PCR analysis a, complete umbel; b, whole buds; c, dissected tepals; d, anthers; e, pistils and ovaries. (B) qRT-PCR expression patterns of PGHF32 genes in floral tissues.
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Figure 5: qRT-PCR analysis of transcripts encoding 1-SST, 6G-FFT, and Cwinv type enzymes in different floral tissues of A. tequilana. (A) Example of flower buds and floral tissues used for qRT-PCR analysis a, complete umbel; b, whole buds; c, dissected tepals; d, anthers; e, pistils and ovaries. (B) qRT-PCR expression patterns of PGHF32 genes in floral tissues.

Mentions: High levels of expression of PGHF32 members in floral tissue have not been documented previously for any Agave species. The in silico expression data however show that many of the isoforms identified for Agave PGHF32 enzymes are highly expressed in floral tissue in all three species. In order to confirm these observations, qRT-PCR analysis was carried out for Atq1-SST-1, Atq6G-FFT-1, Atq6G-FFT-2, and AtqCwinv-1 in immature floral buds (Figure 5A) of A. tequilana samples not used to obtain transcriptome data. As shown in Figure 5B, the fructosyltransferase encoding genes show higher expression in at least one of the floral tissue types in relation to leaf tissue supporting the results from the in silico data. AtqCwinv-1 was expressed at a low level and no significant difference was observed between floral and leaf tissue in this experiment.


New insights into plant glycoside hydrolase family 32 in Agave species.

Avila de Dios E, Gomez Vargas AD, Damián Santos ML, Simpson J - Front Plant Sci (2015)

qRT-PCR analysis of transcripts encoding 1-SST, 6G-FFT, and Cwinv type enzymes in different floral tissues of A. tequilana. (A) Example of flower buds and floral tissues used for qRT-PCR analysis a, complete umbel; b, whole buds; c, dissected tepals; d, anthers; e, pistils and ovaries. (B) qRT-PCR expression patterns of PGHF32 genes in floral tissues.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4524927&req=5

Figure 5: qRT-PCR analysis of transcripts encoding 1-SST, 6G-FFT, and Cwinv type enzymes in different floral tissues of A. tequilana. (A) Example of flower buds and floral tissues used for qRT-PCR analysis a, complete umbel; b, whole buds; c, dissected tepals; d, anthers; e, pistils and ovaries. (B) qRT-PCR expression patterns of PGHF32 genes in floral tissues.
Mentions: High levels of expression of PGHF32 members in floral tissue have not been documented previously for any Agave species. The in silico expression data however show that many of the isoforms identified for Agave PGHF32 enzymes are highly expressed in floral tissue in all three species. In order to confirm these observations, qRT-PCR analysis was carried out for Atq1-SST-1, Atq6G-FFT-1, Atq6G-FFT-2, and AtqCwinv-1 in immature floral buds (Figure 5A) of A. tequilana samples not used to obtain transcriptome data. As shown in Figure 5B, the fructosyltransferase encoding genes show higher expression in at least one of the floral tissue types in relation to leaf tissue supporting the results from the in silico data. AtqCwinv-1 was expressed at a low level and no significant difference was observed between floral and leaf tissue in this experiment.

Bottom Line: Notably however neither 1-FFT nor 6-SFT encoding cDNAs were identified.In silico analysis revealed that distinct isoforms for certain enzymes found in a single species, showed different levels and tissue specific patterns of expression whereas in other cases expression patterns were conserved both within the species and between different species.Thin layer chromatography confirmed the presence of fructans with degree of polymerization (DP) greater than DP three in both immature buds and fully opened flowers also obtained from A. tequilana.

View Article: PubMed Central - PubMed

Affiliation: Department of Genetic Engineering, Centro de Investigación y Estudios Avanzados Irapuato, Mexico.

ABSTRACT
In order to optimize the use of agaves for commercial applications, an understanding of fructan metabolism in these species at the molecular and genetic level is essential. Based on transcriptome data, this report describes the identification and molecular characterization of cDNAs and deduced amino acid sequences for genes encoding fructosyltransferases, invertases and fructan exohydrolases (FEH) (enzymes belonging to plant glycoside hydrolase family 32) from four different agave species (A. tequilana, A. deserti, A. victoriae-reginae, and A. striata). Conserved amino acid sequences and a hypervariable domain allowed classification of distinct isoforms for each enzyme type. Notably however neither 1-FFT nor 6-SFT encoding cDNAs were identified. In silico analysis revealed that distinct isoforms for certain enzymes found in a single species, showed different levels and tissue specific patterns of expression whereas in other cases expression patterns were conserved both within the species and between different species. Relatively high levels of in silico expression for specific isoforms of both invertases and fructosyltransferases were observed in floral tissues in comparison to vegetative tissues such as leaves and stems and this pattern was confirmed by Quantitative Real Time PCR using RNA obtained from floral and leaf tissue of A. tequilana. Thin layer chromatography confirmed the presence of fructans with degree of polymerization (DP) greater than DP three in both immature buds and fully opened flowers also obtained from A. tequilana.

No MeSH data available.