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Isolation and Characterization of a Novel Pathogenesis-Related Protein Gene (GmPRP) with Induced Expression in Soybean (Glycine max) during Infection with Phytophthora sojae.

Jiang L, Wu J, Fan S, Li W, Dong L, Cheng Q, Xu P, Zhang S - PLoS ONE (2015)

Bottom Line: GmPRP was localized in the cell plasma membrane and cytoplasm.Recombinant GmPRP protein exhibited ribonuclease activity and significant inhibition of hyphal growth of P. sojae 1 in vitro.Overexpression of the GmPRP gene in T2 transgenic tobacco and T2 soybean plants resulted in enhanced resistance to Phytophthora nicotianae (P. nicotianae) and P. sojae race 1, respectively.

View Article: PubMed Central - PubMed

Affiliation: Soybean Research Institute, Key Laboratory of Soybean Biology of Chinese Education Ministry, Northeast Agricultural University, Harbin, 150030, Heilongjiang, People's Republic of China.

ABSTRACT
Pathogenesis-related proteins (PR proteins) play crucial roles in the plant defense system. A novel PRP gene was isolated from highly resistant soybean infected with Phytophthora sojae (P. sojae) and was named GmPRP (GenBank accession number: KM506762). The amino acid sequences of GmPRP showed identities of 74%, 73%, 72% and 69% with PRP proteins from Vitis vinifera, Populus trichocarpa, Citrus sinensis and Theobroma cacao, respectively. Quantitative real-time reverse transcription PCR (qRT-PCR) data showed that the expression of GmPRP was highest in roots, followed by the stems and leaves. GmPRP expression was upregulated in soybean leaves infected with P. sojae. Similarly, GmPRP expression also responded to defense/stress signaling molecules, including salicylic acid (SA), ethylene (ET), abscisic acid (ABA) and jasmonic acid (JA). GmPRP was localized in the cell plasma membrane and cytoplasm. Recombinant GmPRP protein exhibited ribonuclease activity and significant inhibition of hyphal growth of P. sojae 1 in vitro. Overexpression of the GmPRP gene in T2 transgenic tobacco and T2 soybean plants resulted in enhanced resistance to Phytophthora nicotianae (P. nicotianae) and P. sojae race 1, respectively. These results indicated that the GmPRP protein played an important role in the defense of soybean against P. sojae infection.

No MeSH data available.


Related in: MedlinePlus

Expression of GmPRP gene in the leaves of transgenic tobacco and soybean plants.(A) qRT-PCR determining the relative bundance of GmPRP (line T2-3 and T2-5) in the transgenic tobacco plants. The non-transgenic tobacco plants only were used as a control. (B) qRT-PCR determining the relative bundance of GmPRP (line T2-3 and T2-5) in the transgenic soybean plants. The non-transgenic soybean plants only were used as a control. (C) Genomic Southern hybridization analysis confirming stable integration and expression of GmPRP in young fully expanded leaves of transgenic soybean plants (CK, wild-type untransformed soybean control, line S2-1 and S2-2 independently transformed T2 transgenic events). All data represent the mean values of three replications.
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pone.0129932.g006: Expression of GmPRP gene in the leaves of transgenic tobacco and soybean plants.(A) qRT-PCR determining the relative bundance of GmPRP (line T2-3 and T2-5) in the transgenic tobacco plants. The non-transgenic tobacco plants only were used as a control. (B) qRT-PCR determining the relative bundance of GmPRP (line T2-3 and T2-5) in the transgenic soybean plants. The non-transgenic soybean plants only were used as a control. (C) Genomic Southern hybridization analysis confirming stable integration and expression of GmPRP in young fully expanded leaves of transgenic soybean plants (CK, wild-type untransformed soybean control, line S2-1 and S2-2 independently transformed T2 transgenic events). All data represent the mean values of three replications.

Mentions: To confirm transgene insertion in transgenic tobacco (T2) and soybean plants (T2), the CTAB method was used to extract genomic DNA from young, fully expanded leaves. Finally, 10 independently transformed T2 transgenic tobacco plants (numbered from T2-1 to T2-10) and 6 independently transformed T2 transgenic soybean plants (numbered from S2-1 to S2-6) were identified by PCR. qRT-PCR of two tobacco transgenic lines (T2-3, T2-5) containing GmPRP and two soybean transgenic lines (S2-1, S2-2) containing GmPRP showed that GmPRP expression was significantly higher than non-transgenic tobacco and soybean plants (Fig 6A and 6B). To determine the copy number of GmPRP in the genome of T2 transgenic soybean, Southern hybridization analysis was performed. Two independently transformed T2 transgenic soybean plants (S2-1 and S2-2) were detected to have three and four copies, respectively (Fig 6C). These results suggest that the GmPRP gene was transformed successfully into soybean plants.


Isolation and Characterization of a Novel Pathogenesis-Related Protein Gene (GmPRP) with Induced Expression in Soybean (Glycine max) during Infection with Phytophthora sojae.

Jiang L, Wu J, Fan S, Li W, Dong L, Cheng Q, Xu P, Zhang S - PLoS ONE (2015)

Expression of GmPRP gene in the leaves of transgenic tobacco and soybean plants.(A) qRT-PCR determining the relative bundance of GmPRP (line T2-3 and T2-5) in the transgenic tobacco plants. The non-transgenic tobacco plants only were used as a control. (B) qRT-PCR determining the relative bundance of GmPRP (line T2-3 and T2-5) in the transgenic soybean plants. The non-transgenic soybean plants only were used as a control. (C) Genomic Southern hybridization analysis confirming stable integration and expression of GmPRP in young fully expanded leaves of transgenic soybean plants (CK, wild-type untransformed soybean control, line S2-1 and S2-2 independently transformed T2 transgenic events). All data represent the mean values of three replications.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4482714&req=5

pone.0129932.g006: Expression of GmPRP gene in the leaves of transgenic tobacco and soybean plants.(A) qRT-PCR determining the relative bundance of GmPRP (line T2-3 and T2-5) in the transgenic tobacco plants. The non-transgenic tobacco plants only were used as a control. (B) qRT-PCR determining the relative bundance of GmPRP (line T2-3 and T2-5) in the transgenic soybean plants. The non-transgenic soybean plants only were used as a control. (C) Genomic Southern hybridization analysis confirming stable integration and expression of GmPRP in young fully expanded leaves of transgenic soybean plants (CK, wild-type untransformed soybean control, line S2-1 and S2-2 independently transformed T2 transgenic events). All data represent the mean values of three replications.
Mentions: To confirm transgene insertion in transgenic tobacco (T2) and soybean plants (T2), the CTAB method was used to extract genomic DNA from young, fully expanded leaves. Finally, 10 independently transformed T2 transgenic tobacco plants (numbered from T2-1 to T2-10) and 6 independently transformed T2 transgenic soybean plants (numbered from S2-1 to S2-6) were identified by PCR. qRT-PCR of two tobacco transgenic lines (T2-3, T2-5) containing GmPRP and two soybean transgenic lines (S2-1, S2-2) containing GmPRP showed that GmPRP expression was significantly higher than non-transgenic tobacco and soybean plants (Fig 6A and 6B). To determine the copy number of GmPRP in the genome of T2 transgenic soybean, Southern hybridization analysis was performed. Two independently transformed T2 transgenic soybean plants (S2-1 and S2-2) were detected to have three and four copies, respectively (Fig 6C). These results suggest that the GmPRP gene was transformed successfully into soybean plants.

Bottom Line: GmPRP was localized in the cell plasma membrane and cytoplasm.Recombinant GmPRP protein exhibited ribonuclease activity and significant inhibition of hyphal growth of P. sojae 1 in vitro.Overexpression of the GmPRP gene in T2 transgenic tobacco and T2 soybean plants resulted in enhanced resistance to Phytophthora nicotianae (P. nicotianae) and P. sojae race 1, respectively.

View Article: PubMed Central - PubMed

Affiliation: Soybean Research Institute, Key Laboratory of Soybean Biology of Chinese Education Ministry, Northeast Agricultural University, Harbin, 150030, Heilongjiang, People's Republic of China.

ABSTRACT
Pathogenesis-related proteins (PR proteins) play crucial roles in the plant defense system. A novel PRP gene was isolated from highly resistant soybean infected with Phytophthora sojae (P. sojae) and was named GmPRP (GenBank accession number: KM506762). The amino acid sequences of GmPRP showed identities of 74%, 73%, 72% and 69% with PRP proteins from Vitis vinifera, Populus trichocarpa, Citrus sinensis and Theobroma cacao, respectively. Quantitative real-time reverse transcription PCR (qRT-PCR) data showed that the expression of GmPRP was highest in roots, followed by the stems and leaves. GmPRP expression was upregulated in soybean leaves infected with P. sojae. Similarly, GmPRP expression also responded to defense/stress signaling molecules, including salicylic acid (SA), ethylene (ET), abscisic acid (ABA) and jasmonic acid (JA). GmPRP was localized in the cell plasma membrane and cytoplasm. Recombinant GmPRP protein exhibited ribonuclease activity and significant inhibition of hyphal growth of P. sojae 1 in vitro. Overexpression of the GmPRP gene in T2 transgenic tobacco and T2 soybean plants resulted in enhanced resistance to Phytophthora nicotianae (P. nicotianae) and P. sojae race 1, respectively. These results indicated that the GmPRP protein played an important role in the defense of soybean against P. sojae infection.

No MeSH data available.


Related in: MedlinePlus