We extracted high-molecular-weight (>50 kbp) DNA from the patient's peripheral blood cells and subsequently labeled the DNA at specific sequence motifs using the Blood and Cell Culture DNA Isolation Kit and NLRS DNA Labeling Kit (Bionano Genomics 80004 and 80001) and nicking enzymes Nt.BspQI and Nb.BssSI (New England Biolabs R0644 and R0681, respectively) as described in respective protocols. The labeled genomic DNA was linearized in nanochannel arrays in a Saphyr chip 1.2 (Bionano Genomics 20319). Single molecules were imaged and digitized using the Saphyr instrument. The output data were used for assembly into genome maps for structural variant detection with the Bionano Access software version 1.4.3.
Blood and cell culture dna isolation kit
Manufactured by Bionano Genomics
The Blood and Cell Culture DNA Isolation Kit is a laboratory equipment product designed to extract and purify DNA from blood and cell culture samples. The kit provides the necessary reagents and protocols to efficiently isolate high-quality genomic DNA for various downstream applications.
Automatically generated - may contain errors
4 protocols using blood and cell culture dna isolation kit
1
High-Molecular-Weight DNA Isolation and Labeling for Structural Variant Detection
2
Lipo863B Genome Optical Mapping
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The genomic DNA from Lipo863B was used for BioNano optical mapping. gDNA was extracted using BioNano Blood and Cell Culture DNA Isolation Kit (80004). Next, the homogenized gDNA was directly labeled using DLE enzyme (BioNano, 80005) according to the BioNano Prep Direct Label and Stain (DLS)-30206 protocol. The labeled and stained genomic DNA was then loaded onto a Saphyr chip for optical data collection.
3
Nanopore Sequencing of Lipo863B gDNA
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The genomic DNA from Lipo863B was used for Nanopore sequencing. gDNA was isolated using BioNano Blood and Cell Culture DNA Isolation Kit (80004). Then, the homogenized gDNA was sheared by pipetting and nanopore library was made using NEBNext Companion Module for Oxford Nanopore Technologies Ligation Sequencing Kit (E7180S) and Nanopore Ligation Sequencing Kit (SQK-LSK110). One microgram of library was loaded per GridION Flow Cell (R9.4.1).
4
Bionano Genomics DNA Extraction and Saphyr Analysis
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DNA was isolated from blood of a six-month-old male FHM using the Blood and Cell Culture DNA Isolation Kit (Bionano Genomics, San Diego, CA). Nucleated fish blood was embedded in agarose plugs for titration in volumes of 1 µL, 3 µL, 8 µL, 20 µL, and 50 µL. A proteinase K digestion was performed, followed by additional washes and agarose digestion. The agarose plugs were sent to the McDonnell Genome Institute (MGI) Washington University for processing and analysis.
From this point, the DNA was drop dialyzed and allowed to equilibrate at room temperature for two days. The DNA samples were assessed for quantity and quality using a Qubit dsDNA BR Assay kit and CHEF gel. A 750ng aliquot of DNA was labeled and stained following the 'Bionano Prep Direct Label and Stain'(DLS) protocol. Once stained, the DNA was quantified using a Qubit dsDNA HS Assay kit and run on the Saphyr chip. During the Saphyr run, the DNA were processed through a series of micro and nanochannels to prevent the linear DNA from folding back on itself. Once the DNA is linearized into the nanochannels, the data were imaged and captured by the Saphyr instrument.
From this point, the DNA was drop dialyzed and allowed to equilibrate at room temperature for two days. The DNA samples were assessed for quantity and quality using a Qubit dsDNA BR Assay kit and CHEF gel. A 750ng aliquot of DNA was labeled and stained following the 'Bionano Prep Direct Label and Stain'(DLS) protocol. Once stained, the DNA was quantified using a Qubit dsDNA HS Assay kit and run on the Saphyr chip. During the Saphyr run, the DNA were processed through a series of micro and nanochannels to prevent the linear DNA from folding back on itself. Once the DNA is linearized into the nanochannels, the data were imaged and captured by the Saphyr instrument.
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