The 16S V2-V3 Library Preparation Kit for Illumina consists of the reagents and components required for library preparation of the 16S V2-V3 amplicon libraries to be used for next-generation sequencing on Illumina platforms. All molecular reagents including primers, enzyme mixes, indexes, and buffers are provided. Instructions for PCR clean up with the AMPure XP Magnetic Beads (supplied by customer) are also included for rapid purification of nucleic acid products generated at two steps of the workflow. The library prep workflow could be used for purified DNA inputs from different sources including stool, soil, water, saliva, plant, urine, skin swab, vaginal swab, cheek swab, nasal swab, plasma/serum, tongue swab, gum swab, and others.
The 16S V2-V3 Library Preparation Kit for Illumina has a streamlined procedure that reduces the handling time such that the library prep procedure can be completed in approximately 4 hours (see diagram below). Input DNA is first subjected to targeted PCR to amplify the V2-V3 region of the DNA encoding 16S rRNA. The post-PCR reaction is then cleaned up using AMPure XP beads. Dual index primers are then added using a limited-cycle PCR. The indexed amplicons flanked by 5′ and 3′ barcoded adaptors are then cleaned using AMPure XP beads. The libraries are then ready for quantification, pooling and sequencing.
Storage Conditions and Product Stability Norgen’s 16S V2-V3 Library Prep Kit for Illumina is shipped as one kit box (for the 24 prep kit) or two sub-component kits (for the 96 prep kit). All kits should be stored at -20°C upon arrival.
All kit components should remain stable for at least 1 year when stored at the specified storage conditions.
Step
Component
Cat. 70300 (24 preps)
Cat. 70310, 70320, 70330, 70340 (96 preps)
Amplicon PCR (PCR 1)
MGX Master Mix
330 µL
1,320 µL
16S V2-V3 Primer Mix
70 µL
280 µL
Index PCR (PCR 2)
Indexing Master Mix
660 µL
2 x 1,320 µL
N7xx Index Primer
50 µL
50 µL
S5xx Index Primer
70 µL
70 µL
PCR Clean-Up
Resuspension Buffer
2 x 1,250 µL
2 x 5,000 µL
Nuclease-free water
1,250 µL
1 x 6,000 µL
Other Products
Endonucleases DNA-specific
Product Info
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Product Info
OverView
HL-dsDNase is especially developed to remove contaminating genomic DNA from RNA preparations. Figure 1 shows that HL-dsDNase can reduce 50 ng of human gDNA to levels non-detectable by qPCR. In figure 2, a human total RNA sample was treated with HL-dsDNase and analysed on the Bio-Rad Experion™ System. The results indicate that HL-dsDNase has minimal impact on RNA quality and quantity.
HL-dsDNase is an engineered version of dsDNase that is rapidly and completely inactivated by incubation for 5 minutes at 58°C with 1 mM DTT at pH 8.0 or above. Chemical inactivation in downstream compatible RT or PCR buffers allows milder heat inactivation and, in some cases, skipping heat inactivation altogether. This makes HL-dsDNase very useful for removal of DNA from RNA preparations since the enzyme may be inactivated using various strategies with reduced risk of auto-degradation of RNA in the presence of magnesium, making HL-dsDNase an ideal enzyme when working with small volumens of RNA.
HL-dsDNase is also an excellent choice for removal of unwanted external DNA from samples prior to analysis of nucleic acids protected by biological membranes, e.g., bacteria, viruses, and sperm. Especially if using downstream analysis methods that might be affected by host cell DNA, as metagenomic sequencing and STR-profiling. The easy inactivation of HL-dsDNase makes it a fast and efficient alternative to methods as differential extraction, where sample material is often lost.
Easily heat-inactivated by very moderate heat treatment
High specific activity
Useful for removal of DNA from RNA preps
Properties
Document
HL-dsDNase is especially developed to remove contaminating genomic DNA from RNA preparations. Figure 1 shows that HL-dsDNase can reduce 50 ng of human gDNA to levels non-detectable by qPCR. In figure 2, a human total RNA sample was treated with HL-dsDNase and analysed on the Bio-Rad Experion™ System. The results indicate that HL-dsDNase has minimal impact on RNA quality and quantity.
Magnetic bead nucleic acid purification technology uses nano or micron superparamagnetic material as the matrix, generally black ferric oxide or yellowish brown ferricoxide as the magnetic material. The surface of bead is coated with appropriate functional groups, which can adsorb nucleic acid. Magnetic beads commonly used for nucleic acids, containing carboxyl groups, hydroxyl groups, or silicon groups. Silicon-based magnetic beads are the most common, and its principle of adsorbing nucleic acid is consistent with the classical glass milk purification technology or glass fiber filter membrane purification method. Magpure particle is a kind of polydisperse fast speed silica magnetic beads. The core is ferricoxide, accounting for 50%, and the surface coating is silica, accounting for 50%. The product can be used for plasmid extraction, gel DNA recovery, product purification, genomic DNA and RNA extraction, and viral nucleic acid extraction.
Details
Specifications
Features
Specifications
Concentration
40 mg/ml
Appearance
Suspension of dark brown particles
Surface functional group
Si-OH, Silanol
Dispersibility
Monodisperse,spherical
Particle size
1.0-1.5 μm
Preservation conditions
Room Temperature, valid for up to 2 years.It is recommended to store in 2-8°C to prevent microbial growth.
Magnetic response speed
~30 seconds
Settling velocity
>3 minutes
High salt mediated binding
>2M guanidine isothiocyanate, DNA recovery up to 80%
Alcohol mediated binding
2M guanidine hydrochloride / isopropanol (30%), and the recovery of DNA / RNA was as high as 85%
PEG8000 mediated binding
The recovery of DNA/RNA was up to 85%
DNase/RNase
Not detected
DNA residue
Not detected
Recommended application
Genomic DNA extraction, RNA extraction, viral nucleic acid extraction, circulating DNA isolation
Principle
Highsalt mediated binding: in the solution containing 2-4M guanidine isothiocyanate, Magpure particles can selectively recover DNA molecules, and impurities such as protein polysaccharides are not adsorbed.
Alcohol mediated binding: in the solution containing guanidine salt and alcohol (>25%), Magpure particles can selectively recover DNA/RNA molecules, and proteins and other impurities are not adsorbed.
After biological samples are treated with digestive solution or lysis Buffer, DNA/RNA is released from cells, organelles and protein complexes (ribosomes and nucleosomes) into reagents. After Magpure particles and binding solution are added, DNA/RNA is adsorbed to the surface of Magpure particles to form DNA/RNA bead complex. Under the action of the magnetic field, the magnetic beads are separated and collected, and the impurities such as protein are removed with the waste liquid. After two or three steps of further cleaning, the DNA/RNA magnetic bead complex is resuspended in sterilized water or TE buffer, and the DNA/RNA falls off from the surface of the magnetic beads, so as to achieve the purpose of purification.
gDNA/RNA Isolation from Blood, Tissue, Plant, Swab, Spots, Stool, Soil and etc.Viral DNA/RNA IsolationAgarose Gel DNA Purification
DNA/RNA Isolation from low nucleic acid content samplesPlasmid IsolationDNA/RNA Clean Up
Circulating DNA IsolationViral Nucleic acid IsolationgDNA Isolation FFPE DNA/RNA Isolation
Plasmid extractiongel DNA recoverygenomicDNA/RNA extraction viral nucleic acid extractionCirculating DNA extraction
DNA/RNA Clean Up and concentrationDNA/RNA Isolation from low nucleic acid content samplesResearch immuno assays
The MagPure magnetic-particle technology combines the speed and efficiency of silica-based DNA purification with the convenient handling of magnetic particles. DNA binds to the silica surface of the magnetic particles in the presence of a chaotropic salt. DNA bound to the particles is then efficiently washed, considerably improving the purity of DNA. High-quality DNA is eluted. The automated purification procedure completely removes enzymes, nucleotides, and other contaminants and inhibitors. Purified DNA is suitable for direct use in downstream applications, such as sequencing and microarray analysis.
Document
Magnetic bead nucleic acid purification technology uses nano or micron superparamagnetic material as the matrix, generally black ferric oxide or yellowish brown ferricoxide as the magnetic material. The surface of bead is coated with appropriate functional groups, which can adsorb nucleic acid. Magnetic beads commonly used for nucleic acids, containing carboxyl groups, hydroxyl groups, or silicon groups. Silicon-based magnetic beads are the most common, and its principle of adsorbing nucleic acid is consistent with the classical glass milk purification technology or glass fiber filter membrane purification method. Magpure particle is a kind of polydisperse fast speed silica magnetic beads. The core is ferricoxide, accounting for 50%, and the surface coating is silica, accounting for 50%. The product can be used for plasmid extraction, gel DNA recovery, product purification, genomic DNA and RNA extraction, and viral nucleic acid extraction.
This product is suitable for extracting total viral nucleic acid from cell-free/low-content cell biological samples such as body fluids, serums, plasma, soaking solutions, tissue homogenate supernatant, and culture supernatant. The Purified DNA/RNA is used for RT-PCR and PCR detection.
Details
Specifications
Features
Specifications
Main Functions
Extract viral DNA/RNA from200μl samples by magnetic beads
Applications
RT-PCR，PCR，NGS
Products
Viral DNA / RNA, body cell DNA / RNA
Purification method
Polydisperse magnetic beads
Purification technology
Magnetic beads technolog
Process method
Manual or automatic
Sample type
Sample amount
200μl
Adaptive instrument
Nucleic acid extractor, pipetting workstation
Yield
Elution volume
Time per run
Liquid carrying volume per column
Binding yield of column
Principle
This product is based on the purification method of high binding magnetic particles. The sample is lysed and digested under the action of lysate and Protease. DNA/RNA is released into the lysate. After adding magnetic particles and binding solution, DNA/RNA will be adsorbed on the surface of magnetic particles, and impurities such as proteins will be removed without adsorption. The adsorbed particles were washed with washing solution to remove proteins and impurities, washed with ethanol to remove salts, and finally DNA/RNA was eluted by Nuclease Free Water.
Advantages
Fast – several samples can be extracted in 40 minutes by column method
High quality – high purity total RNA / DNA can be directly used in various sensitive downstream applications
Safe – no phenol chloroform extraction required
Sensitive – DNA/RNA can be recovered at the level of PG
Kit Contents
Contents
IVD5412
Purification Times
200 Preps
MagPure Particles N
5 ml
PK/Carrier RNA
50 mg
Protease Dissolve Buffer Blue
5 ml
Buffer MLB
120 ml
Buffer MW1
53 ml
RNase Free Water
30 ml
Storage and Stability
This kit is shipped and stored at room temperature and is valid for 12 months.
Experiment Data
Document
This product is suitable for extracting total viral nucleic acid from cell-free/low-content cell biological samples such as body fluids, serums, plasma, soaking solutions, tissue homogenate supernatant, and culture supernatant. The Purified DNA/RNA is used for RT-PCR and PCR detection.
