Guidelines DiBio Sequencing service
Booking and delivery
Samples must be booked via email specifying:
- organism
- RNAseq or DNAseq
- library protocol
- number of samples
- million reads/sample
Shipping condition: Dry Ice
Ship to:
NGS Service
Biology Department
University of Padova
Via U.Bassi 58/B 35121-Padova
Tubes must contain only DNA or RNA and should be free of any contamination.
Delivery: by appointment (0498276160 or ngs.cribi@unipd.it).
RNAseq Illumina mRNA Prep protocol
● 2 ug of Total RNA 100 ng/ul (Qubit) in nuclease-free water;
● high quality Total RNA RIN>7 as measured at the Agilent TapeStation;
● minimize DNA contamination (if possible verify the DNA quantity at the Qubit fluorometer);
● use 0.5ml or 0.2ml tubes with single caps labeled with the codes provided by the facility. Samples codes will be generated after receiving an excel file from the user with sample names.
RNAseq Lexogen 3’mRNA protocol
● annotated genome is required since the sequences are generated at the 3'-UTR of the gene;
● 2 ug of Total RNA 150 ng/ul (quantified at the Qubit) in nuclease-free water;
● high quality Total RNA RIN>7 as measured at the Agilent TapeStation;
● minimize DNA contamination (if possible verify the DNA quantity at the Qubit fluorometer);
● use 0.2 ml strip tubes with single caps and write codes (Identification Numbers) provided by the facility. Samples codes will be generated after receiving an excel file
from the user with sample names.
● this sequencing approach is fitting also samples partially degraded (please contact us for information)
DNAseq Illumina protocol
● 200 ng of genomic DNA 7-10 ng/ul (Qubit) in nuclease-free water;
● the protocol is optimized for DNA with 260/280 spectrophotometer absorbance ratio values of 1.8-2.0 and 260/230 absorbance ratio values of 2.0-2.2;
● use 0.2ml strip tubes with single caps with codes (Identification Numbers) provided by the facility. Samples codes will be generated after receiving an excel file from the user with sample names.
Samples and data storage
● samples will be stored at -80° until the end of the project only upon request by the user at the time of booking. The user have to recover the samples at the facility
within 2 weeks;
● sequencing data will be stored for 6 months in our servers and then they will be removed; it is in charge of the user to take care of long term storage, and it is highly
reccommended to download and save immediately the fastq files.
For RNAseq ribodepletion process (rRNA removal), 16S sequencing, bioinformatic analysis, and any further information please contact us: ngs.cribi@unipd.it 0498276160
Guidelines RNA/DNA quantity/quality service
Booking and delivery
Fill in the ‘Utilizzatore’ field specifying:
- user mail address
- number of samples
- sample names
Sample must be delivered by appointment (0498276160 or ngs.cribi@unipd.it).
They have to contain only DNA or RNA and be free of any contamination.
Agilent Tapestation protocol
Assay Sizing range Quantitative range
RNA 25-500 ng/ul
RNA HS 0.5-10 ng/ul
D1000 35-1000 bp 0.1-50 ng/ul
D5000 100-5000 bp 0.1-50 ng/ul
genomic DNA 200 to >6000 bp 10-100 ng/l
● use 0.2 ml strip tubes with single caps;
● please provide 2 ul/sample.
Thermofisher Qubit protocol
● prepare samples following assay instructions;
Assay Initial sample concentration Quantitation range
DNA HS 0.005-120 ng/ul 0.1-120 ng
RNA HS 0.2-200 ng/ul 4-200 ng
RNA BR 0.5-1200 ng/ul 10-1200 ng
● use 0.2ml strip tubes with single cap;
● please provide at least 5ul/sample.
Thermofisher Nanodrop protocol
● Users have to register the number of samples online before coming to the instrument for the measures
● the user guide is next to the instrument
● use your tips and micropipettes
● leave the workstation clean and tidy
● empty the bin
● turn off the instrument
● for any clarification please contact the personnel
Samples storage
● samples will be discarded after sending the results, unless otherwise indicated at the
time of booking
For any further information please contact us: ngs.cribi@unipd.it 049 8276160
