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Insights into transcriptional characteristics and homoeolog expression bias of embryo and endosperm in developing grain through mRNA-Seq and Iso-Seq

ABSTRACT: To analyse the regulation of transcripts in grain embryo and endosperm during development, we performed RNA-Seq for wheat from 14 and 25 day post anthesis (DPA). And long-read sequencing for mixed whole grains from 14 and 25 DPA was employed to obtain full-length transcripts. A series of differentially expressed genes and tissues-specific genes of embryo and endosperm were identified. Moreover, 4351, 4641, 4516 and 4453 genes with A, B and D homoeoloci were detected in the four tissues. These provide specific gene pools of embryo and endosperm and homoeolog expression bias model in a large scale, which provides new insights into the molecular physiology of wheat.

ORGANISM(S): Triticum aestivum

PROVIDER: GSE118474 | GEO | 2018/08/14

REPOSITORIES: GEO

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Project description:A microarray analysis of wheat grain hardness For both the Heron and Falcon NIL, whole wheat heads were harvested at 6, 15 and 25 days post anthesis (DPA) from glass house grown material and stored at -80 OC. For the other wheat cultivars analysed, heads at approximately 9-10 DPA were used. Eight to ten developing caryopses at the same age and morphological stage of development were selected from a head to represent a sample. Seed collected from a different head represented a replicate sample. Total RNA was isolated from the whole seed following the method of Higgins et al. (1976) with the following modification to remove starch. After the lithium chloride precipitation, the RNA pellet was resuspended in 250 ?l water. Then 3.5 ?l of 3 M sodium acetate (pH 5.3) and 125 ?l ethanol were added and the sample was centrifuged for 10 minutes at 4 OC. The supernatant was transferred to a fresh tube and the RNA was ethanol-precipitated by the addition of 21.5 ?l of 3 M sodium acetate (pH 5.3) and 375 ?l ethanol. The recovery of RNA varied from 0.06-0.5 ?g RNA per mg of tissue and this was dependent on the developmental stage of the seeds used for extraction of the RNA. For the labelling procedure, 50 ?g of total RNA was used for both the Cy3 and Cy5 dyes (Amersham Pharmacia, UK), following the two-step labelling method of Schenk et al. (2000). The Cy3 control (hard wheat) and Cy5 sample (soft wheat) were swapped among the replicate experiments to minimise any bias in cDNA incorporation and photo-bleaching of the fluorescent dyes. The pre-hybridisation of the microarray slides, hybridisation of the target cDNA and subsequent washing of the slides to remove unbound target was performed as per the supplied protocol for the CMT-GAPSTM coated microscope slides (Corning USA). The slides were scanned with a GenePix 4000A microarray scanner (Axon Instruments, Union, CA, USA). The features were analysed using the GenePix Pro 4 software and unsatisfactorily segmented features were either manually adjusted or discarded to ensure the integrity of the data obtained.

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Insights into transcriptional characteristics and homoeolog expression bias of embryo and endosperm in developing grain through mRNA-Seq and Iso-Seq

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