The NGS Cell Free DNA Library Prep Kit (illumina and MGI Platforms) was developed for the construction of high quality cell-free DNA (cfDNA) libraries using 1 ng to 50 ng of cell-free DNA as input. The kit has a simple work flow and a fast procedure. Multiplexing of the cell free DNA library is possible based on the index type.
NGS Cell Free DNA Library Prep Kit Workflow
The main source of cell-free DNA (cfDNA) is derived from apoptotic hematopoietic cells in blood and found in the plasma. The length of the cfDNA is about 150-200 bp in length. Circulating tumor DNA (ctDNA) derived from malignant tumors is a part of cfDNA. Both cfDNA and ctDNA can be used as a noninvasive biomarker since it offers a better approach in comparison to invasive tissue biopsies.
NGS has been used for cfDNA and ctDNA sequencing in the field of liquid biopsy as it provides a whole genome level of molecular profiling. One of the hurdles for cfDNA sequencing is the difficulty of library preparation from the limited amount of cfDNA obtained from plasma. Our cell-free NGS kit makes it easy to get enough libraries from limited input in just 1.5 hours.
Three index types are available for the NGS Cell Free DNA Library Prep Kit of the illumina platform:
Non-index (Cat.# 30029): Libraries do not have index.
Index(Cat.# 30031): Each of our index primers contains a unique 6-base index sequence that can be used for sample identification. Total 48 library multiplexing is possible. Index information can be downloaded here.
Unique dual index (Cat.# 30033): Cell-free DNA library multiplexing up to 96 samples is possible with the unique dual indexes. We have developed a Four-Base Difference Index System. The system have at least 4 bases different from each other in the 8 bases index length. The primers effectively minimize sequencing errors such as mis-assignment, index hopping, index contamination etc. Index information can be downloaded here.
Indexes are available for the MGI platform kits (Cat.# 34031).
Kit advantages:
Comparison of library conversion efficiency with different samples under the same condition.
Comparison of library yield with different samples under the same condition.
The NGS Cell Free DNA Library Prep Kit (illumina and MGI Platforms) was developed for the construction of high quality cell-free DNA (cfDNA) libraries using 1 ng to 50 ng of cell-free DNA as input. The kit has a simple work flow and a fast procedure. Multiplexing of the cell free DNA library is possible based on the index type.
Name of Product
Lactobacillus brevis
Catalog Number
MGLBr Z
Short Info
GenLine discovers beer germs in a timely and precise manner
Method/Platform
PCR
Range/Assay Sensivity
10^4 -10^5 cfu/mL
Brief Instructions
Storage
-15°C to -20°C
Components
PCR-Polymerase Master-Mix, PCR-Primermix, Positive Kit-Control, PCR-Water
Test Principle
The technological basis for the GenLine tests is the polymerase chain reaction (PCR) combined with lateral flow Tests.
Name of Product
Lactobacillus brevis
Catalog Number
MGLBr Z
Short Info
GenLine discovers beer germs in a timely and precise manner
Method/Platform
PCR
Range/Assay Sensivity
10^4 -10^5 cfu/mL
Test Principle
The technological basis for the GenLine tests is the polymerase chain reaction (PCR) combined with lateral flow Tests.
Endonucleases DNA-specific, dsDNase
Double-Strand Specific dsDNase (dsDNase) is ideal for fast and effective removal of contaminating DNA from PCR master mixes.
Taq polymerases are commonly contaminated by bacterial DNA. This is a problem in PCR based bacterial typing and detection as it might cause false positive results. The unique properties of dsDNase make it suited for removal of contaminating DNA from PCR master mixes prior to addition of DNA template.
In figure 1, a PCR master mix was treated with different amounts of dsDNase before performing a qPCR to measure the contaminating bacterial DNA in the master mix. ArcticZymes dsDNase effectively removed contaminating DNA below known levels of the assay detection limits.
The dsDNase from Arctic shrimp (Pandalus borealis) is recombinantly produced in Pichia pastoris. It cleaves phosphodiester linkages in DNA to yield oligonucleotides with 5’-phosphate and 3’-hydroxyl termini.
The specific activity is estimated to be 30 times higher than that of bovine DNase I. In the presence of magnesium as only divalent cation and using oligos as a substrate, the activity towards dsDNA is 5000-fold higher than towards ssDNA.
The unique double strand-specificity allows specific degradation of dsDNA while leaving shorter ssDNA as primers and probes essentially intact. Easy inactivation by moderate heat (65°C) allows addition of DNA intended for analysis directly after removal of contaminating DNA.
Figure 1. The dsDNase effectively removes contaminated DNA
The dsDNase effectively removes contaminated DNA:
A PCR master mix was preincubated with various concentrations of dsDNase. After treatment, no DNA was amplified in non-template controls.
Nucleic acid specificity has been tested towards double- and single-stranded DNA and RNA oligonucleotides. The specificity of dsDNase towards the substrate has been measured using 15-mer oligonucleotides with FAM at 5′ and DarkQuencher® 3′ (Eurogentec). The fluorescence is proportional to enzyme activity. Assay conditions: 25 mM Tris pH 7.5, 5 mM MgCl2, and 2 μM oligonucleotide.
Substrate Relative Activity
dsDNA 100%
ssDNA <0.03%
dsRNA <0.01%
ssRNA <0.01%
Double-Strand Specific dsDNase (dsDNase) is ideal for fast and effective removal of contaminating DNA from PCR master mixes.
Taq polymerases are commonly contaminated by bacterial DNA. This is a problem in PCR based bacterial typing and detection as it might cause false positive results. The unique properties of dsDNase make it suited for removal of contaminating DNA from PCR master mixes prior to addition of DNA template.
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