Description
Material provided: data
Unit definition: 24 samples – 1M reads per sample
Short Description: Since 2019 the spread of SARS-CoV2 virus has quickly evolved in a global pandemic. Whole genome sequencing has become an important tool to understand how the virus genome evolves and to discover emerging variants important to global community health. This hybridization capture method generates overlapping targets to cover the 29.9 kb viral genome using probes designed against the NCBI Reference sequence NC_045512. The reconstruction of complete Sars-Cov2 genome would allow variant designation by detecting known and novel strains and support phylogenetic and prevalence studies.
Ideal for the discovery and the profiling of the Sars-Cov-2 known and emerging variants.
Benefits of generate Sars-Cov2 hybridization capture libraries directly from total RNA to characterize coronavirus variants:
- The preparation of hybridization capture libraries optimizes Sars-Cov2 genome sequencing by targeting short specific regions of the genome.
- Whole genome sequencing allows the identification and the characterization of known and potential novel viral sequences.
The Experiment and the Product
The sequencing analysis service Sars-Cov-2 Illumina sequencing (Ref-SKU: S.1.13.2.IT.149.40) based on target sequencing, includes the following steps for each sample:
- Reverse transcription of total RNA into single strand cDNA ;
- Preparation of cDNA libraries ;
- Target Enrichment ;
- Post capture PCR amplification ;
- Library Validation by Fragment Analyzer ;
- Normalization to 4nM for cluster generation ;
- Sequencing : 2×150 PE MiSeq/NextSeq550
Figure 1. Fragment analyzer trace for the target-based library validation step
– END-USER RESPONSIBILITIES Biological material to be provided by the end-users and described in the dedicated request form.
– RNA INPUT RECOMMENDATIONS
For the Target-based library preparation, the Polo d’Innovazione Genomica Genetica Biologia, follows the library preparation protocol from Twist resources. The kit protocol has been optimized for Total RNA.
Please, provide at least 100ng of Total RNA. Minimum volume: 50 µl/molecular grade water.
– REMARKS:
- If the provided sample doesn’t comply with the above requested specifications for “RNA INPUT RECOMMENDATIONS”, the customer will be contacted for a re-sampling.
- In case of failure of the sequencing run, the service will be repeated.
– SAMPLE SHIPPING
- Safe-lock 1.5 – 2 ml tubes properly capped to be used to avoid accidental evaporation or sample contamination during shipping.
- Samples to be labelled correctly.
- Correct temperature must be ensured during the shipment to avoid sample degradation.
- Complete the enclosed Sample Submission form and return together with the samples to: orders@pologgb.com.
SUPPLIER RESPONSIBILITIES:
– STORAGE OF BIOLOGICAL SAMPLE
Polo d’Innovazione Genomica Genetica Biologia will store the analysed samples only for 1 month after delivery. Extended storage time is available for a maximum of 4 months and must be requested in advance of the experiments in the service request application.
– STORAGE OF DATA
Polo d’Innovazione Genomica Genetica Biologia waives any responsibility for the storage and backup of the supplied project and sequencing data after delivery to the end-user. Polo d’Innovazione Genomica Genetica Biologia will store the project data only for 1 month after delivery. After expiration of this period, all data are removed from our servers and archived (unless otherwise agreed with the end-user). Your data might be re-imported at an extra cost within the period of 1 year after delivery of the final data. After 1 year, all project data will be deleted permanently from our systems. Extended data storage is available on request at an extra cost and must be requested at project start.
– DELIVERY OF RESULTS
The sequencing results will be sent according to the Modalities of Transfer, with a QC Report PDF File 90 days upon samples delivery by the User.
Modalities of transfer:
- by “WeTransfer” platform for data of up to 2Gb
- by “MegaSync” platform for data ranging from 2Gb to 15Gb
- by HD for data over 15 Gb
Please note that the cost of HD driver is not included. Where requested, the end-user must provide the HD.
For further details on the description of the experiment please consult the following links:
- Creating cDNA Libraries using Twist Library Preparation EF Kit 2.0 for ssRNA Virus Detection
- Twist Target Enrichment Fast Hybridization Protocol
For more information, please contact us.
For technical assistance, contact Polo GGB Technical Support:
Email: techsupport@pologgb.com
Telephone: 0039 0577 381310