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High throughput expression profiling of induction of microspore reprogramming to embryogenesis in Brassica napus L. [dataset 1]

ABSTRACT: High throughput expression profiling of induction of microspore reprogramming to embryogenesis in Brassica napus L. [dataset 1]

PROVIDER: PRJNA1186138 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	SRR31350278_1.fastq.gz	Fastqsanger.gz
	SRR31350278_2.fastq.gz	Fastqsanger.gz
	SRR31350279_1.fastq.gz	Fastqsanger.gz
	SRR31350279_2.fastq.gz	Fastqsanger.gz
	SRR31350280_1.fastq.gz	Fastqsanger.gz

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High throughput expression profiling of induction of microspore reprogramming to embryogenesis in Brassica napus L. [dataset 1]

Project description:Microspore embryogenesis is an in vitro system in which haploid microspores (precursor celsl of pollen grains) are reprogrammed towards embryogenesis by the application of a heat stress (HS) treatment. This process is of great interest in plant breeding as it enables accelerated production of double-haploid plants. The induction of in vitro microspore reprogramming comprises a complex chain of events that still remain unknown. The process can be induced at the responsive developmental stage of vacuolated microspore. In order to evaluate the transcriptional changes acompaying microspore embryogenesis induction in Brassica napus, an expression profiling through high throughput RNA sequencing, and a pipeline for bioinformatic analysis was performed. The analyses have been implemented for two samples: (1) vacuolated microspores (VM), before stress treatment, initial stage of the microspore culture, and (2) stress-treated microspores, 4 days after the application of the HS, when proembryos (PE) are formed; PE correspond to the first morphological sign of embryogenesis induction. The results obtained would provide valuable information about this complex process.

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The transcriptional landscape of microspore embryogenesis induction in barley

Project description:The effectiveness of microspore embryogenesis and plant formation from in vitro cultivated microspores is determined by a complex network of internal and environmental factors. Plant breeding and genetic engineering widely use haploids/doubled haploids (DHs) derived from in vitro-cultured microspores, but the mechanism of ME induction remains poorly defined. Here, RNA-sequencing was used to characterize the transcriptional landscapes of four early stages of ME in two barley cultivars (Golden Promise and Igri) that are contrastingly responsive in the microspore-derived plant formation. Our experimental setup revealed fundamental regulatory networks, key marker genes of ME (628 stage-specific markers), and 160 candidate genes crucial for genotype-dependent responsiveness/resistance to ME. We found that transcription factors probably give rise to embryo-like structure and callus formation at stage III and determine their further development. Our high-resolution temporal transcriptome atlas provides an important resource for future functional studies aiming at unravelling the genetic control of microspore transition.

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Effect of HDAC inhibitor TSA on haploid embryogenesis [dataset 1]

Project description:Haploid embryos can be induced from cultured immature pollen following a stress treatment. In Brassica napus, application of the histone/lysine deacetylase (HDAC/KDAC) inhibitor trichostatin A (TSA) to pollen cultures enhances the production of differentiated embryos and embryogenic callus when applied together with heat stress (Li et al., 2014). To identify genes associated with the induction of B. napus haploid embryogenesis, we compared the transcriptomes of untreated pollen cultures and pollen cultures treated with either heat-stress or heat-stress plus TSA.

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Project description:Haploid embryos can be induced from cultured immature pollen following stress treatment. In Brassica napus, the application of the histone/lysine deacetylase (HDAC/KDAC) inhibitor trichostatin A (TSA) to pollen cultures enhances the production of differentiated embryos and embryogenic callus when applied together with heat stress (Li et al., 2014). To identify genes associated with the induction of haploid embryogenesis and to investigate which genes may be responsible for TSA-induced lipid and starch accumulation in the embryogenic structures, we compared the transcriptomes of pollen cultures treated with heat stress and 0.05 µM TSA to those of untreated pollen cultures, both at two days.

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Isolation of an embryogenic line from non-embryogenic Brassica napus cv. Westar through microspore embryogenesis.

Project description:Brassica napus cultivar Westar is non-embryogenic under all standard protocols for induction of microspore embryogenesis; however, the rare embryos produced in Westar microspore cultures, induced with added brassinosteroids, were found to develop into heritably stable embryogenic lines after chromosome doubling. One of the Westar-derived doubled haploid (DH) lines, DH-2, produced up to 30% the number of embryos as the highly embryogenic B. napus line, Topas DH4079. Expression analysis of marker genes for embryogenesis in Westar and the derived DH-2 line, using real-time reverse transcription-PCR, revealed that the timely expression of embryogenesis-related genes such as LEAFY COTYLEDON1 (LEC1), LEC2, ABSCISIC ACID INSENSITIVE3, and BABY BOOM1, and an accompanying down-regulation of pollen-related transcripts, were associated with commitment to embryo development in Brassica microspores. Microarray comparisons of 7 d cultures of Westar and Westar DH-2, using a B. napus seed-focused cDNA array (10 642 unigenes), identified highly expressed genes related to protein synthesis, translation, and response to stimulus (Gene Ontology) in the embryogenic DH-2 microspore-derived cell cultures. In contrast, transcripts for pollen-expressed genes were predominant in the recalcitrant Westar microspores. Besides being embryogenic, DH-2 plants showed alterations in morphology and architecture as compared with Westar, for example epinastic leaves, non-abscised petals, pale flower colour, and longer lateral branches. Auxin, cytokinin, and abscisic acid (ABA) profiles in young leaves, mature leaves, and inflorescences of Westar and DH-2 revealed no significant differences that could account for the alterations in embryogenic potential or phenotype. Various mechanisms accounting for the increased capacity for embryogenesis in Westar-derived DH lines are considered.

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Effect of HDAC inhibitor TSA on haploid embryogenesis [dataset 1]

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