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Expression of Pennisetum glaucum Eukaryotic Translational Initiation Factor 4A ( PgeIF4A ) Confers Improved Drought, Salinity, and Oxidative Stress Tolerance in Groundnut

View Article: PubMed Central - PubMed

ABSTRACT

Eukaryotic translational initiation factor 4A belong to family of helicases, involved in multifunctional activities during stress and non-stress conditions. The eIF4A gene was isolated and cloned from semi-arid cereal crop of Pennisetum glaucum. In present study, the PgeIF4A gene was expressed under the regulation of stress inducible Arabidopsis rd29A promoter in groundnut (cv JL-24) with bar as a selectable marker. The de-embryonated cotyledons were infected with Agrobacterium tumefaciens (LBA4404) carrying rd29A:PgeIF4A construct and generated high frequency of multiple shoots in phosphinothricin medium. Twenty- four T0 plants showed integration of both nos-bar and rd29A-PgeIF4A gene cassettes in genome with expected amplification products of 429 and 654 bps, respectively. Transgene copy number integration was observed in five T0 transgenic plants through Southern blot analysis. Predicted Mendelian ratio of segregation (3:1) was noted in transgenic plants at T1 generation. The T2 homozygous lines (L1-5, L8-2, and L16-2) expressing PgeIF4A gene were exhibited superior growth performance with respect to phenotypic parameters like shoot length, tap root length, and lateral root formation under simulated drought and salinity stresses compared to the wild type. In addition, the chlorophyll retention was found to be higher in these plants compared to the control plants. The quantitative real time—PCR results confirmed higher expression of PgeIF4A gene in L1-5, L8-3, and L16-2 plants imposed with drought/salt stress. Further, the salt stress tolerance was associated with increase in oxidative stress markers, such as superoxide dismutase accumulation, reactive oxygen species scavenging, and membrane stability in transgenic plants. Taken together our results confirmed that the PgeIF4A gene expressing transgenic groundnut plants exhibited better adaptation to stress conditions.

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Related in: MedlinePlus

Expression of PgeIF4A transcript in transgenic groundnut (T2) lines. (A) Semi quantitative RT-PCR showing transcript expression pattern of PgeIF4A and G6PD (Glucose 6 phosphate 1 dehydrogenase) in control and transgenic lines grown under mannitol (drought) and NaCl (salinity) stress conditions. (B) qRT-PCR expression analysis of transgenic plants under un treated (control) and treated (drought and salinity) conditions. The Ct-values of the samples were normalized with G6PD house keeping gene. Lane C: non-transgenic, Lane: L1-5, L8-3, and L16-2 represented T2 transgenic lines.
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Figure 7: Expression of PgeIF4A transcript in transgenic groundnut (T2) lines. (A) Semi quantitative RT-PCR showing transcript expression pattern of PgeIF4A and G6PD (Glucose 6 phosphate 1 dehydrogenase) in control and transgenic lines grown under mannitol (drought) and NaCl (salinity) stress conditions. (B) qRT-PCR expression analysis of transgenic plants under un treated (control) and treated (drought and salinity) conditions. The Ct-values of the samples were normalized with G6PD house keeping gene. Lane C: non-transgenic, Lane: L1-5, L8-3, and L16-2 represented T2 transgenic lines.

Mentions: The expression of PgeIF4A transcript was confirmed by semi quantitative PCR and qRT-PCR. The PgeIF4A transcript accumulated in homozygous T2 lines (L1-5, L8-3, and L16-2) grown under stress (mannitol and NaCl) conditions (Figure 7A). The qRT-PCR results showed that the PgeIF4A transcript could be detected in all three transgenic plants grown under normal and stress conditions (Figure 7B). The quantitative regulation of PgeIF4A in response to drought/salt stress was higher than the plants grown under normal growth conditions. The transgene expression in drought and salinity treated plants was about 4–5-folds higher than the transgenic lines grown under normal conditions (Figure 7B). It indicated that the transgene expression is positively correlated upon stress imposition.


Expression of Pennisetum glaucum Eukaryotic Translational Initiation Factor 4A ( PgeIF4A ) Confers Improved Drought, Salinity, and Oxidative Stress Tolerance in Groundnut
Expression of PgeIF4A transcript in transgenic groundnut (T2) lines. (A) Semi quantitative RT-PCR showing transcript expression pattern of PgeIF4A and G6PD (Glucose 6 phosphate 1 dehydrogenase) in control and transgenic lines grown under mannitol (drought) and NaCl (salinity) stress conditions. (B) qRT-PCR expression analysis of transgenic plants under un treated (control) and treated (drought and salinity) conditions. The Ct-values of the samples were normalized with G6PD house keeping gene. Lane C: non-transgenic, Lane: L1-5, L8-3, and L16-2 represented T2 transgenic lines.
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Related In: Results  -  Collection

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Figure 7: Expression of PgeIF4A transcript in transgenic groundnut (T2) lines. (A) Semi quantitative RT-PCR showing transcript expression pattern of PgeIF4A and G6PD (Glucose 6 phosphate 1 dehydrogenase) in control and transgenic lines grown under mannitol (drought) and NaCl (salinity) stress conditions. (B) qRT-PCR expression analysis of transgenic plants under un treated (control) and treated (drought and salinity) conditions. The Ct-values of the samples were normalized with G6PD house keeping gene. Lane C: non-transgenic, Lane: L1-5, L8-3, and L16-2 represented T2 transgenic lines.
Mentions: The expression of PgeIF4A transcript was confirmed by semi quantitative PCR and qRT-PCR. The PgeIF4A transcript accumulated in homozygous T2 lines (L1-5, L8-3, and L16-2) grown under stress (mannitol and NaCl) conditions (Figure 7A). The qRT-PCR results showed that the PgeIF4A transcript could be detected in all three transgenic plants grown under normal and stress conditions (Figure 7B). The quantitative regulation of PgeIF4A in response to drought/salt stress was higher than the plants grown under normal growth conditions. The transgene expression in drought and salinity treated plants was about 4–5-folds higher than the transgenic lines grown under normal conditions (Figure 7B). It indicated that the transgene expression is positively correlated upon stress imposition.

View Article: PubMed Central - PubMed

ABSTRACT

Eukaryotic translational initiation factor 4A belong to family of helicases, involved in multifunctional activities during stress and non-stress conditions. The eIF4A gene was isolated and cloned from semi-arid cereal crop of Pennisetum glaucum. In present study, the PgeIF4A gene was expressed under the regulation of stress inducible Arabidopsis rd29A promoter in groundnut (cv JL-24) with bar as a selectable marker. The de-embryonated cotyledons were infected with Agrobacterium tumefaciens (LBA4404) carrying rd29A:PgeIF4A construct and generated high frequency of multiple shoots in phosphinothricin medium. Twenty- four T0 plants showed integration of both nos-bar and rd29A-PgeIF4A gene cassettes in genome with expected amplification products of 429 and 654 bps, respectively. Transgene copy number integration was observed in five T0 transgenic plants through Southern blot analysis. Predicted Mendelian ratio of segregation (3:1) was noted in transgenic plants at T1 generation. The T2 homozygous lines (L1-5, L8-2, and L16-2) expressing PgeIF4A gene were exhibited superior growth performance with respect to phenotypic parameters like shoot length, tap root length, and lateral root formation under simulated drought and salinity stresses compared to the wild type. In addition, the chlorophyll retention was found to be higher in these plants compared to the control plants. The quantitative real time—PCR results confirmed higher expression of PgeIF4A gene in L1-5, L8-3, and L16-2 plants imposed with drought/salt stress. Further, the salt stress tolerance was associated with increase in oxidative stress markers, such as superoxide dismutase accumulation, reactive oxygen species scavenging, and membrane stability in transgenic plants. Taken together our results confirmed that the PgeIF4A gene expressing transgenic groundnut plants exhibited better adaptation to stress conditions.

No MeSH data available.


Related in: MedlinePlus