Analysis of Differences in DNA Methylation in Organically and  Conventionally Grown Potato Tubers (Solanum tuberosum)
Analysis of Differences in DNA Methylation in Organically and  Conventionally Grown Potato Tubers (Solanum tuberosum)
For the study DNA from potato tubers was used. The potato tubers (Solanum tuberosum var. Desirée) we harvested from the DOK-field trial (D: bio-dynamic, O: bio-organic, K: con-ventional) in Therwil, Switzerland of Agroscope, the Swiss Centre of Excellence for Agri-cultural Research and the Research Institute of Organic Agriculture (Foschungsinstitut für biologischen Landbau - FIBL). The DOK is one of the largest and the longest lasting long-term field trial of its kind, initiated in 1978. The samples have been kindly provided by Agroscope. Fully developed tubers (according BBCH Scale 909) were collected at harvest time (Hack et al. 1993) from potato plants (Solanum tuberosum cv. Desiree) in four replicates of the bio-organic cropping system (organic) and the conventional cropping system (conventional) from four years (2001, 2009, 2011, 2012). The complete tubers formed a representative sample of the plot per biological replicate.  Library preparation and sequencing was performed at Novogene Inc. (Novogene Co., Ltd. Novogene Biotech Co., Ltd. Building 301, Zone A10 Jiuxianqiao North Road, Chao-yang District, Beijing). After transfer of samples integrity, quantity and quality of DNA was controlled. Afterwards the DNA has been sheared using Covaris S220 to fragments of 200-400 bp and further processed according the library preparation protocol including terminal repair, A-ligation and ligation of sequencing adapters. HiSeq X Ten (Illumina Inc., 9885 Towne Centre Drive. San Diego, CA 92121 U.S.A.) sequencing, with paired-end 150 bp, including multiplexing of other, non-bisulfite treated, samples has been applied to account for the unbalanced base ratio in WGBS libraries. The effective concentration of the final library has been >2 nM. A Lambda Phi X spike-in with 0.5% of DNA weight has been applied as conversion standard. The demultiplexed raw data has been received. The potato tubers (Solanum tuberosum var. Desirée) we harvested from the DOK-field trial (D: bio-dynamic, O: bio-organic, K: conventional) in Therwil, Switzerland of Agroscope, the Swiss Centre of Excellence for Agricultural Research and the Research Institute of Organic Agriculture (Foschungsinstitut für biologischen Landbau - FIBL). (D: bio-dynamic, O: bio-organic, K: conventional)  Whole-Genome-Bisulfite Sequencing (WGBS) procedure using a Post-Bisulfite Adapt-er-Tagging protocol (PBAT) to account for low amounts of DNA using the EZ DNA Meth-ylation Gold-TM Kit, Zymo Research. DNA concentration after bisulfite conversion was quantified using Qubit and q-PCR. column-based DNeasy kit according to manufacturer (Qiagen) protocol, including a first phenol purification step with 100 mg of the ground potato sample. Integrity of DNA was tested by gel electrophoresis and Ethidiumbromid staining  a Whole-Genome-Bisulfite Sequencing (WGBS) procedure using a Post-Bisulfite Adapt-er-Tagging protocol (PBAT) to account for low amounts of DNA using the EZ DNA Meth-ylation Gold-TM Kit, Zymo Research. DNA concentration after bisulfite conversion was quantified using Qubit and q-PCR.