Overview

• Protocol optimized for DNA isolated from a diversity of samples including stool, soil, water, saliva, plant, urine, skin, and more
• Simple and quick workflow: library could be prepared in less than 5 hours
• Component of Norgen’s metagenomics workflow
• A single NGS run can be prepared with up to 384 unique dual-index libraries

Sample type purification kit guide 

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.

Details

Supporting Data

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Minimum amount of starting material:	2.5 µL of DNA (5 ng/µL)
Time to complete library preparation:	4 hours

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

Introduction

The HiPure Mini system provides a fast, simple, and cost-effective plasmid DNA miniprep method for routine molecular biology laboratory applications. HiPure Plasmid Mini Kits use silica membrane technology to eliminate the cumbersome steps associated with loose resins or slurries. Plasmid DNA purified with Mini Kits is immediately ready for use. Phenol extraction and ethanol precipitation are not required, and high quality plasmid DNA is eluted in a small volume of Tris buffer or water.

Details

Specifications

Features	Specifications
Main Functions	Isolation up to 70μg plasmid DNA from 5-15ml bacterial culture
Applications	Enzyme digestion, sequencing, PCR, cloning, etc.
Purification method	Mini spin column
Purification technology	Silica technology
Process method	Manual (centrifugation or vacuum)
Sample type	Conventional plasmid, plasmid less than 30KB
Sample amount	High copy plasmid: 1-5ml culture mediumLow copy plasmid : 5-10ml culture medium
Yield	20-80µg
Elution volume	≥75μl
Time per run	Complete 1-24 samples in 30 minutes
Liquid carrying volume per column	800µl
Binding yield of column	70µg

Principle

The HiPure Plasmid procedure is based on alkaline lysis of bacterial cells followed by adsorption of DNA onto silica in the presence of high salt. The unique silica membrane used in the kit completely replaces glass or silica slurries for plasmid DNA minipreps. The procedure consists of 3 basic steps: Preparation and clearing of a bacterial lysate by alkaline method，then transfer the supernatant to column to bind DNA. After washing proteins and other impurities, nucleic acid was finally eluted with low-salt buffer (10mm Tris, pH9.0, 0.5mm EDTA).

Advantages

• High purity – purified plasmid can be directly used in sequencing, enzyme digestion and PCR, etc.
• Fast – it takes only 2 minutes to obtain supernatant by optimized solution (10 minutes for other brands)
• High yield – up to 70µg plasmid can be binded in one column

Kit Contents

Contents	P100202	P100203
Purification Times	50 Preps	250 Preps
RNase A	5 mg	20 mg
Buffer P1	30 ml	140 ml
Buffer P2	30 ml	140 ml
Buffer P3	40 ml	200 ml
Buffer PW1	30 ml	140 ml
Buffer PW2*	12 ml	50 ml
Elution Buffer	15 ml	30 ml
HiPure DNA Mini Columns III	50	250 ml
2 ml Collection Tubes	50	250 ml

Storage and Stability

The kit components can be stored dry at room temperature (15-25°C) and are stable for at least 18 months under these conditions. If any precipitates form in the buffers, warm at 37℃ to dissolve. After addition of RNase A，Buffer P1 is stable for 6 months when stored at 2-8°C.

Purchase Guide

Name	CAT NO	Features	Sampleamount	Elutionvolume	Yield
HiPure Plasmid Mini Kit	P1001	Classic, Regular application	1-5ml LB	30-50μl	5–35µg/5ml
HiPure Plasmid Mini Kit II	P1002	Classic, Regular application	5-15ml LB	75-100μl	20-80µg/15ml
HiPure Plasmid Plus 96 Kit	P1006	Classic, Regular application	1-1.5ml LB	70-150μl	1-15μg/1ml
HiPure Fastfilter Plasmid Midi Kit	P1013	Fast	30-50ml LB	0.3-0.8ml	50–250µg/50ml
HiPure Fastfilter Plasmid Maxi Kit	P1014	Fast	100-200ml LB	0.5-1.5ml	0.4–1mg/200ml
Low endotoxin/Endotoxin-free Plasmid
HiPure Plasmid EF Maxi Kit	P1156	Classic method, General for all kinds of vectors	100-200ml LB	≥0.7ml	200–1500µg
HiPure Plasmid EF Mini Kit	P1154	Recommend for low copy vector, Thoroughly remove RNA	5-15ml LB	≥60μl	10–80µg
HiPure Plasmid EF 96 Kit	P1157	Recommend for low copy vector, Thoroughly remove RNA	1-5ml LBx96	≥75μl	30µg
HiPure Plasmid EF Midi Kit	P1231	Recommend for High copy vector, Low elution volume, High concentration	25-50ml LB	≥100μl	10-500μg
HiPure Plasmid EF Mega Kit	P1116	Recommend for High copy vector(>3μg/ml)	500ml LB	≥2ml	1-10mg

For any technical problems or customized products, please contact us.

F&Q about Endotoxin-free Plasmid Extraction Kit — P1156 ←click here

The HiPure Mini system provides a fast, simple, and cost-effective plasmid DNA miniprep method for routine molecular biology laboratory applications. HiPure Plasmid Mini Kits use silica membrane technology to eliminate the cumbersome steps associated with loose resins or slurries. Plasmid DNA purified with Mini Kits is immediately ready for use. Phenol extraction and ethanol precipitation are not required, and high quality plasmid DNA is eluted in a small volume of Tris buffer or water.

Description 

The ExcelTaq™ 2X Fast Q-PCR Master Mix (SYBR, ROX) is a ready-to-use reagent with all the essential components for quantitative real-time PCR (qPCR) except primers and templates. The master mix contains a highly stable hot-start Taq polymerase in an optimized buffer with dsDNA specific SYBR green fluorescent dye. Consequently, the ExcelTaq™ 2X Fast Q-PCR Master Mix (SYBR, ROX) features high stability during storage, even at 37°C for weeks. In addition to high sensitivity and signal intensity, the ExcelTaq™ 2X Fast Q-PCR Master Mix (SYBR, ROX) performs a low background/ high specificity qPCR results, as well as a better compatibility with fast PCR program. With inert smart blue contrast dye, the ExcelTaq™ 2X Fast Q-PCR Master Mix (SYBR, ROX) is ready-to-use and greatly reduces pipetting errors, while largely improving the reproducibility of the process. The ExcelTaq™ 2X Fast Q-PCR Master Mix (SYBR, ROX) also includes ROX reference dye for normalizing the fluorescent reporter signal in real-time quantitative PCR. This master mix allows sensitive, precise amplification, real-time tracking of the amplification process, and simultaneous quantification for targeted DNA molecules.

Features

• High Stability
• Fast Hot Start
• High Sensitivity
• Low Background
• Suitable for Fast Program 
• Smart Blue Contrast Dye 
• With ROX Reference Dye

Storage

Protect from light.
Aliquot to avoid multiple freeze-thaw cycles. 
-20°C for 12 months

The ExcelTaq™ 2X Fast Q-PCR Master Mix (SYBR, ROX) is a ready-to-use reagent with all the essential components for quantitative real-time PCR (qPCR) except primers and templates. The master mix contains a highly stable hot-start Taq polymerase in an optimized buffer with dsDNA specific SYBR green fluorescent dye. Consequently, the ExcelTaq™ 2X Fast Q-PCR Master Mix (SYBR, ROX) features high stability during storage, even at 37°C for weeks. In addition to high sensitivity and signal intensity, the ExcelTaq™ 2X Fast Q-PCR Master Mix (SYBR, ROX) performs a low background/ high specificity qPCR results, as well as a better compatibility with fast PCR program. With inert smart blue contrast dye, the ExcelTaq™ 2X Fast Q-PCR Master Mix (SYBR, ROX) is ready-to-use and greatly reduces pipetting errors, while largely improving the reproducibility of the process. The ExcelTaq™ 2X Fast Q-PCR Master Mix (SYBR, ROX) also includes ROX reference dye for normalizing the fluorescent reporter signal in real-time quantitative PCR. This master mix allows sensitive, precise amplification, real-time tracking of the amplification process, and simultaneous quantification for targeted DNA molecules.