NGS DNA Library Prep Kit (illumina and MGI Platforms)
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The kit was developed for construction of high quality libraries for next generation sequencing (illumina and MGI Platforms). The kit needs double strand DNA fragments (blunt and/or sticky) as input DNA for NGS library preparation, and is compatible with DNA fragments generated from both enzymatic methods and physical methods (sonication, nebulization etc.). Library multiplexing is possible with different types of indexes.
Detail
The kit was developed for construction of high quality libraries for next generation sequencing (illumina and MGI Platforms). The kit needs double strand DNA fragments (blunt and/or sticky) as input DNA for NGS library preparation, and is compatible with DNA fragments generated from both enzymatic methods and physical methods (sonication, nebulization etc.). Library multiplexing is possible with different types of indexes.
The kit was optimized for next generation sequencing (NGS) library preparation with different types of samples. Most of DNA library preparation requires the ligation of sheared DNA fragments to library adaptors and the DNA library preparation is closely related to the quality of NGS data. With BioDynami’s unique DNA library preparation technologies, the fast and simple kit allows high quality NGS library preparation to be completed in 1.5 hours with only 10 minutes of hands-on time.
Some genomic regions are very difficult to be covered evenly and usually result in very low coverage rate or gap in these regions.
Typical difficult regions are: • with high GC contents • have secondary structures: mainly due to repeat sequences • the worst cases: have both high GC contents and repeated sequences.
Example: human TERT gene is one of the most difficult regions as shown above. NGS data showed that BioDynami kit has the best performance to cover the extremely difficult human TERT gene region.
Limited GC bias across whole genome
Three index types are available for the illumina platform kits:
Non-index (illumina Cat.# 30009): Libraries do not have index.
Index (illumina Cat.# 30021): A unique barcode sequence with 6 bases has been included in each of the index primers. RNA Sequencing library multiplexing is possible with up to 48 samples. Index information can be downloaded here.
Unique dual index (illumina Cat.# 30023): RNA Sequencing library multiplexing up to 96 samples is possible with the unique dual indexes. We have developed a 4-Base Difference Index System. The system can generate indexes with at least 4 bases different from others in the 8-base indexing region. the unique dual indexing primers identify sequencing errors such as index hopping, mis-assignment, and de-multiplexing errors. Index information can be downloaded here.
Indexes are available for the MGI platform kits (Cat.# 34021).
Our Blyscan™ Glycosaminoglycan Kit has been a ‘go-to’ Solution for reliable sGAG and Proteoglycan Analysis for many years! Blyscan utilises a dye-binding approach to quantitatively measure sulfated glycosaminoglycans (sGAG) and proteoglycans in cells, tissues and fluids from a wide range of in-vivo and in-vitro sources.
Colorimetric Detection (656nm) (Endpoint)
Understanding Glycosaminoglycans (GAGs) and Proteoglycans
Glycosaminoglycans (GAGs) are a type of negatively charged polysaccharide that play crucial roles in various biological processes. They are composed of repeated disaccharide units, typically of N-acetylated or N-sulfated hexosamine paired with a uronic acid (GlcA or IdoA) or galactose. Sulfate groups can also be added to give sulfated GAGs an overall negative charge that influences cell interactions and also enable binding by our Blyscan dye reagent.
Common examples of GAGs include Chondroitin Sulfate, Dermatan Sulfate, Heparin, Heparan Sulfate, and Keratan Sulfate. Note that Hyaluronic Acid is a non-sulfated GAG and cannot be detected by the Blyscan assay. If you need to measure hyaluronic acid instead, we recommend using our Purple-Jelley kit!
The Role of Glycosaminoglycans in Tissues
GAGs and proteoglycans have essential functions in tissues and organisms, providing biophysical support through scaffolding and maintaining cartilage hydration. They also play a vital role in biochemical processes such as cell adhesion and signalling.
What is the origin of the Blyscan assay name?
Blyscan is an Old English word meaning ‘to shine’ and from which the word ‘blush’, (blushing), may have been derived. This was an appropriate choice as the Blyscan Assay contains a blue dye which ‘blushes’ bright pink when it binds to sulphated glycosaminoglycans!
How does the Blyscan assay work?
Step 1. Blyscan dye reagent contains DMMB dye in an optimised buffer. Addition of Dye reagent to samples containing sGAG results in the formation of a dye/sGAG complex due to a charge interaction between dye and GAG sulfate groups.
Step 2. Over a 30 minute incubation Dye-labelled sGAGs precipitate out of solution and are collected by centrifugation. Following removal of unbound dye, the remaining bound dye is released from the complex by addition of dye dissociation reagent. Released dye is quantified spectrophotometrically.
Step 3. The sGAG content of unknown samples may be quantified by comparison against a calibration curve prepared using a standard of purified Chondroitin-4-sulfate supplied with the kit.
A list of suggested sample types can be found under the ‘Assay Specification‘ tab.
The Blyscan Dye reagent is formulated to miminise binding to other charged sample components such as nucleic acids, a problem with some older dye-based sGAG assays.
Assay range
2.5 – 50µg/ml
Limit of Detection
2.5µg/ml
Detection Method
Colorimetric Detection (656nm) (Endpoint)
Measurements per kit
110 in total (allows a maximum of 48 samples to be run in duplicate alongside a standard curve).
In-vivo: Liquid samples, including fluids such as urine, amniotic or synovial fluid.
In-vitro: Solid samples, such as deposited ECM on 2D/3D culture surfaces.by enzymatic treatment
In-vivo: Liquid samples, Culture media during 2D/3D cell culture.
The assay requires that sulfated polysaccahrides or sGAGs are in a soluble form. A preliminary enzymatic extraction step is required for solid samples (enzyme not supplied with kit).
The assay is not suitable for use with samples containing alginates or that comprise degraded sulfated disaccharide fragments.
Precautions
This kit is designed for research use only. Not for use in diagnostic procedures. Kit requires access to a centrifuge, as well as a spectrophotometer/colorimeter capable of absorbance detection at 656nm. Specific sample preparation protocols may require customer to provide further reagents, consult assay manual for further information.
Blyscan sGAG kit contents:
1. Blyscan Dye Reagent (1x110ml)
2.sGAG Reference Standard (1x5ml, 100µg/ml Bovine tracheal chondroitin 4-sulfate)
3. Dissociation Reagent (1x110ml)
4. Sodium Nitrite (1x15ml)
5. Acetic Acid (1x15ml)
6. Ammonium Sulfamate (1x15ml)
7. 1.5ml micro-centrifuge tubes for dye-labelling reaction.
8. Assay kit manual
NB: Additional reagents may be required for sample preparation prior to assay. Consult manual or contact us for further details.
Document
Our Blyscan™ Glycosaminoglycan Kit has been a ‘go-to’ Solution for reliable sGAG and Proteoglycan Analysis for many years! Blyscan utilises a dye-binding approach to quantitatively measure sulfated glycosaminoglycans (sGAG) and proteoglycans in cells, tissues and fluids from a wide range of in-vivo and in-vitro sources. Colorimetric Detection (656nm) (Endpoin
PACE (PCR Allelic Competitive Extension) genotyping chemistry is a homogeneous, PCR-based allele-specific technology for the analysis of DNA sequence variants, most commonly SNPs (Single Nucleotide Polymorphisms) and Indels (insertion / deletions).
PACE genotyping chemistry is comprised of two parts:
PACE Genotyping Assay: two allele-specific forward primers and one common, reverse primer. The allele-specific forward primers each have different 3′ terminal bases reflecting the target variant, and a unique 5’ tail sequence which is incorporated as part of the fluorescent signal mechanism.
PACE Genotyping Master Mix: containing all remaining components required for PCR and the generation of fluorescent signals. PACE Genotyping Master Mix contains a novel, universal, fluorescent reporting cassette to produce machine-readable fluorescent signals (FAM and HEX) corresponding to the assay genotypes.
When combined with sample DNA, these components create a PACE Genotyping Reaction, as illustrated in the figure below.
We have extensive knowledge and experience in assay design, especially when it comes to allele-specific PCR. PACE Genotyping Assays are available to purchase either Validated and Unvalidated. Validated assays require customer DNA to validate and optimise, for guaranteed performance. Unvalidated assays are designed in silico and supplied untested.
REQUIRED COMPONENTS
qPCR machine or Thermocycler + Fluorescent plate reader
PCR plate or equivalent and appropriate optically clear seal
Template DNA
PCR-grade water
PACE Genotyping Master Mix or PACE 2.0 Genotyping Master Mix
STEPS TO YOUR PACE GENOTYPING ASSAY DESIGN
Place your order on this page. Our support team will contact you by email.
Fill out an Assay Design Template form with all the information we need to process your custom PACE Genotyping Assay order. We will email you a copy of the template when we first contact you, or your can download a copy here.
Using the information you provide us, we will create your PACE Genotyping Assay designs, order the oligos, and send you design sequences.
Once we receive the oligos, we assemble the assay(s) and then ship an aliquot to you (unvalidated) or test on your DNA samples before shipping the aliquot to you (validated).
