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In situ embryo rescue for generation of wide intra- and interspecific hybrids of Panicum virgatum L.

ABSTRACT: Wide crosses have been used for decades as a method for transferring novel genetic material and traits in plant breeding. Historically, many products of wide crosses require tedious and inefficient surgical embryo rescue prior to embryo abortion to recover single plantlets. We have utilized transgenic switchgrass (Panicum virgatum L. cv Alamo) as a pollen donor in conjunction with antibiotic or herbicide selection for recovery of intra-and interspecific F₁ crosses by using developing ovules from the female parent and selecting for embryogenic cultures derived from the in situ immature embryo. Using this approach, several intravarietial crosses were generated between transgenic Alamo and the switchgrass varieties Kanlow, Blackwell and Cave-in-Rock as well as an interspecific cross with Atlantic coastal panicgrass. This procedure selected F₁ embryogenic callus produced from the developing embryo contained within isolated immature ovules. Several clonal plants were successfully regenerated from each cross. Southern blot, PCR, phenotypic analyses and genomic analysis confirmed F₁ hybrids. Using genotyping-by-sequencing shows the hybridization of the recovered plants by determining the ratio of transgressive markers to total compared markers between parents and their potential offspring. The ratio of transgressive markers to total compared markers was significantly lower between parents and their predicted offspring than between parents and offspring unrelated to them. This approach provides the possibility to move useful transgenes into varieties that are recalcitrant to direct transformation which can be optionally segregated thus useful to create new hybrids, as well as recovery of wide crosses that are either difficult or impossible using traditional techniques.

SUBMITTER: Kausch AP

PROVIDER: S-EPMC5095774 | biostudies-literature | 2016 Nov

REPOSITORIES: biostudies-literature

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In situ embryo rescue for generation of wide intra- and interspecific hybrids of Panicum virgatum L.

Kausch Albert P AP Tilelli Michael M Hague Joel J Heffelfinger Christopher C Cunha David D Moreno Maria M Dellaporta Stephen L SL Nelson Kimberly K

Plant biotechnology journal 20160601 11

Wide crosses have been used for decades as a method for transferring novel genetic material and traits in plant breeding. Historically, many products of wide crosses require tedious and inefficient surgical embryo rescue prior to embryo abortion to recover single plantlets. We have utilized transgenic switchgrass (Panicum virgatum L. cv Alamo) as a pollen donor in conjunction with antibiotic or herbicide selection for recovery of intra-and interspecific F<sub>1</sub> crosses by using developing ...[more]

PMID: 27154282

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Project description:BackgroundGlobal warming predictions indicate that temperatures will increase by another 2-6°C by the end of this century. High temperature is a major abiotic stress limiting plant growth and productivity in many areas of the world. Switchgrass (Panicum virgatum L.) is a model herbaceous bioenergy crop, due to its rapid growth rate, reliable biomass yield, minimal requirements of water and nutrients, adaptability to grow on marginal lands and widespread distribution throughout North America. The effect of high temperature on switchgrass physiology, cell wall composition and biomass yields has been reported. However, there is void in the knowledge of the molecular responses to heat stress in switchgrass.ResultsWe conducted long-term heat stress treatment (38°/30°C, day/night, for 50 days) in the switchgrass cultivar Alamo. A significant decrease in the plant height and total biomass was evident in the heat stressed plants compared to controls. Total RNA from control and heat stress samples were used for transcriptome analysis with switchgrass Affymetrix genechips. Following normalization and pre-processing, 5365 probesets were identified as differentially expressed using a 2-fold cutoff. Of these, 2233 probesets (2000 switchgrass unigenes) were up-regulated, and 3132 probesets (2809 unigenes) were down-regulated. Differential expression of 42 randomly selected genes from this list was validated using RT-PCR. Rice orthologs were retrieved for 78.7% of the heat stress responsive switchgrass probesets. Gene ontology (GOs) enrichment analysis using AgriGO program showed that genes related to ATPase regulator, chaperone binding, and protein folding was significantly up-regulated. GOs associated with protein modification, transcription, phosphorus and nitrogen metabolic processes, were significantly down-regulated by heat stress.ConclusionsPlausible connections were identified between the identified GOs, physiological responses and heat response phenotype observed in switchgrass plants. Comparative transcriptome analysis in response to heat stress among four monocots - switchgrass, rice, wheat and maize identified 16 common genes, most of which were associated with protein refolding processes. These core genes will be valuable biomarkers for identifying heat sensitive plant germplasm since they are responsive to both short duration as well as chronic heat stress treatments, and are also expressed in different plant growth stages and tissue types.

Dataset Information

In situ embryo rescue for generation of wide intra- and interspecific hybrids of Panicum virgatum L.

Publications

In situ embryo rescue for generation of wide intra- and interspecific hybrids of Panicum virgatum L.

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