Whole-genome library preparation from WGA DNA

Whole-genome libraries were prepared from approximately 100 ng WGA DNA using NEBNext Ultra II FS Kit following the protocol for inputs ≤100 ng (New England BioLabs, Ipswich, Massachusetts, USA), with the following modifications: the NEBNext Adaptor for Illumina was replaced with the Universal iTru adapter [13 (link)]. The libraries were amplified with iTru i5 and i7 primers [13 (link)] at a the final concentration of 0.5 μM, purified using 1x Sera-Mag Select Beads (Cytiva, Amersham, Little Chalfont, United Kingdom), and resuspended in a final volume of 33 μL of 0.1× TE buffer.

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Nguyen H.D., Ponomareva E., Dadej K., Smith D., Antoun M., van der Lee T.A, & van de Vossenberg B.T. (2024). A target enrichment approach for enhanced recovery of Synchytrium endobioticum nuclear genome sequences. PLOS ONE, 19(2), e0296842.

Publication 2024

NEBNext Ultra II FS Kit Sera-Mag Select Beads

Corresponding Organization : Agriculture and Agri-Food Canada

Other organizations : Canadian Food Inspection Agency, Wageningen University & Research

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Variable analysis

independent variables

Use of NEBNext Ultra II FS Kit for library preparation
Replacement of NEBNext Adaptor for Illumina with Universal iTru adapter
Use of iTru i5 and i7 primers for library amplification at a final concentration of 0.5 μM

dependent variables

Quality of whole-genome libraries prepared

control variables

Amount of WGA DNA used (approximately 100 ng)
Purification of libraries using 1x Sera-Mag Select Beads
Resuspension of libraries in 0.1× TE buffer with a final volume of 33 μL

controls

No positive or negative controls were explicitly mentioned in the provided information.

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