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Sample GSM3926942

Query DataSets for GSM3926942

Status

Public on Jul 07, 2021

Title

CS_EC2

Sample type

SRA

Source name

Embryogenic callus tissue

Organism

Vitis vinifera

Characteristics

cultivar: Cabernet Sauvignon
embryogenesis aptitude: poorly competent
tissue: Embryogenic callus tissue
biological replicate: Replicate 2

Growth protocol

Anthers (A) were cultured on a previously described callus induction medium supplemented with 4.5 µM 2,4-dichlorophenoxyacetic acid (2,4-D) and 8.9 μM 6-benzylaminopurine (BAP) (Gribaudo et al., 2004; Gambino et al., 2007). The cultures were maintained at 26 °C in the dark. Three months after the initiation of the culture, calli were transferred for 1 months to embryo proliferation medium (Gribaudo et al., 2004; Gambino et al., 2007) containing 10 µM 2-Napthoxyacetic acid (NOA), 1 μM BAP, 20 µM Indole-3-acetic acid (IAA) and 2.5% activated charcoal. The frequency of callogenesis and embryogenesis for both cultivars was calculated in two consecutive years at 4 months after the initiation of the culture, and calli were classified as embryogenic (EC) or non-embryogenic (NEC) under a stereomicroscope.

Extracted molecule

total RNA

Extraction protocol

Total RNA was extracted from approximately 100 mg of anthers, 40d calli, EC and NEC. For each biological replicate, the plant material ground in liquid nitrogen using the tissue lyser system (TissueLyser II, Qiagen), was extracted using the Spectrum Plant Total RNA extraction kit (Sigma Aldrich). The integrity and quantity of RNA were checked with a 2100 Bioanalyzer Chip RNA 7500 series II (Agilent, Santa Clara, CA, USA)) and a Nanodrop 2000 spectrophotometer (Thermo Scientific, Wilmington, DE, USA).
The anthers, 40d calli, NEC and EC triplicate samples yielded 24 non-directional cDNA libraries, which were prepared from 2.5 µg of total RNA using the Illumina TruSeq RNA Sample preparation protocol (Illumina Inc., San Diego CA, USA) according to the manufacturer’s instructions.

Library strategy

RNA-Seq

Library source

transcriptomic

Library selection

cDNA

Instrument model

Illumina HiSeq 2000

Data processing

base-calling (Illumina Casava v. 1.8.2)
Illumina quality filtering (custom script)
alignment of reads to the reference genome (TopHat v. 2.0.9)
transcripts assembly and generation of normalized abundance measurements (Cufflinks v. 2.1.1)
Genome_build: Vitis vinifera PN40024 reference genome V1 (assembly 12x). Jaillon O, et al. (2007) The grapevine genome sequence suggests ancestral hexaploidization in major angiosperm phyla. Nature 449: 463-467
Supplementary_files_format_and_content: FPKMs

Submission date

Jul 03, 2019

Last update date

Jul 07, 2021

Contact name

Silvia Dal Santo

E-mail(s)

silvia.dalsanto@univr.it

Organization name

University of Verona

Department

Department of Biotechnology