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 V1-V2 Library Preparation Kit for Illumina consists of the reagents and components required for library preparation of the 16S V1-V2 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 V1-V2 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 V1-V2 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

Figure 1 / 3

Previous

Next

Click for expanded view

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 V1-V2 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. 70100 (24 preps)	Cat. 70110, 70120, 70130, 70140 (96 preps)
Amplicon PCR (PCR 1)	MGX Master Mix	330 µL	1,320 µL
	16S V1-V2 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

MagZol Reagent is a reagent system for the isolation of total RNA from cells and tissues. The reagent, a single-phase solution consisting of phenol and guanidine isothiocyanate, is modification of the single-step RNA isolation method developed by Chomczynski and Sacchi. The sample is homogenized and lysed in MagZol Reagent which maintains the integrity of the RNA, while disrupting and denaturing endogenous RNases and other cellular components. Extraction of the lysate with chloroform further denatures proteins and separates the mixture into an organic and an aqueous phase. RNA remains exclusively in the aqueous phase, and is subsequently recovered by isopropanol.

This method is suitable for small quantities of tissue (<100mg) and cells (<5 X10⁶), and large quantities of tissue (up to 1g) and cells (<10⁸), of human, animal, plant, or bacterial origin. The simplicity of the MagZol Reagent method allows simultaneous processing of a large number of samples. The entire procedure can be completed in one hour. Total RNA prepared in this manner can be used for Northern blot analysis, dot blot hybridization, poly(A) + selection, in vitro translation, RNase protection assay, and molecular cloning. For use in amplification by thermal cycling, treatment of the isolated RNA with RNase-free DNase I is recommended when the two amplimers lie within a single exon.

Details

Specifications

Features	Specifications
Main Functions	Extract RNA from liquid samples by salting out method
Applications	RT-PCR, Northern hybridization, poly (a) enrichment, etc.
Purification technology	Acid phenol guanidine isothiocyanate
Process method	Manual (centrifugation)
Sample type	Various liquid samples
Sample amount	Flexible
Elution volume	Variation with sample size
Time per run	Variation with sample size

Advantages

• Flexible – sample amount can be adjusted according to the demand
• Cost performance -the most economical nucleic acid extraction technology

Storage and Stability

MagZol Reagent should be stored at 2-8°C upon arrival and is stable for at least 24 months under the condition. However, short-term storage (up to 12 weeks) at room temperature (15-25°C) does not affect its performance.

MagZol Reagent is a reagent system for the isolation of total RNA from cells and tissues. The reagent, a single-phase solution consisting of phenol and guanidine isothiocyanate, is modification of the single-step RNA isolation method developed by Chomczynski and Sacchi. The sample is homogenized and lysed in MagZol Reagent which maintains the integrity of the RNA, while disrupting and denaturing endogenous RNases and other cellular components. Extraction of the lysate with chloroform further denatures proteins and separates the mixture into an organic and an aqueous phase. RNA remains exclusively in the aqueous phase, and is subsequently recovered by isopropanol.

Overview

• Isolate high quality, high yield plasmid DNA
• Plasmid DNA is ready for various downstream applications including restriction digestion, bacterial transformation, sequencing and more
• Available in 4 formats: MiniPrep, MiniPrep (Magnetic Bead System), 96-Well MiniPrep (Magnetic Bead System), and MaxiPrep

These kits are designed for the rapid preparation of plasmid DNA from Escherichia coli.

Plasmid MiniPrep Kit

This kit is designed for the rapid preparation of plasmid DNA from small cultures of Escherichia coli using convenient spin columns. The plasmid DNA is preferentially purified from other cellular components such as genomic DNA and RNA. This kit is able to purify plasmids up to 13,000 bp in size, and the typical purification yield is up to 20 μg from 1.5 mL of bacterial culture. Purified DNA is of excellent quality for transformation, restriction enzyme digestion, sequencing and more. Also available in a 96-well format.

Plasmid MiniPrep Kit (Magnetic Bead System and High Throughput Magnetic Bead System)

Norgen’s Plasmid MiniPrep Kit (Magnetic Bead System) is designed for the rapid preparation of plasmid DNA from small batch cultures of Escherichia coli. Norgen’s magnetic beads bind DNA under optimized salt concentrations and release the bound DNA under low salt and slightly alkali conditions. The plasmid DNA is preferentially purified from other cellular components such as genomic DNA and RNA. The purified plasmids are fully digestible with all restriction enzymes tested, and are completely compatible with real-time PCR and NGS.

Norgen’s Plasmid MiniPrep Kit (Magnetic Bead System) is also available in a 96-well (HT) format for high throughput applications. Purification with the 96-well plates can be integrated with a robotic automation system.

Plasmid MaxiPrep Kit

This kit is designed for the rapid spin column preparation of plasmid DNA from up to 100 mL of Escherichia coli cultures. The kit allows for the isolation of plasmid DNA with final endotoxin levels of 0.1 EU/µg of DNA or less. The kit is able to purify plasmids up to 13,000 bp in size, and typical yields from a 100 mL culture for a high copy number plasmid are between 0.4 and 1.0 mg. The purified DNA is fully digestible with all restriction enzymes tested, and is completely compatible with manual or automated sequencing to achieve 95-100% accuracy.

Details

Supporting Data

Figure 1 / 5

Previous

Next

Click for expanded view

Kit Specifications
Column Binding Capacity	1.5 mg
Average Yield from 100 mL Culture*	0.4 – 1.0 mg
Final Endotoxin Levels	< 0.1 EU/µg DNA
Time to Complete 4 Purifications	1.5 hours
Size of Plasmids Purified	Up to 13,000 bp

 * Depends on high-copy DNA and low copy DNA

Storage Conditions and Product Stability
All solutions should be kept tightly sealed and stored at room temperature. The RNase vial should be stored at -20°C upon arrival. Once RNase has been added to the Resuspension Solution AZ the solution should be stored at 4°C. This kit is stable for 1 year after its date of shipment.

Component	Cat. 13300 (50 preps)	Cat. 46400 (250 preps)	Cat. 46500 (4 preps)	Cat. 46600 (20 preps)	Cat. 60300 (50 preps)	Cat. 63000 (192 preps)
Resuspension Solution AZ	12 mL	60 mL	20 mL	100 mL	12 mL	2 x 20 mL
Lysis Buffer N	40 mL	80 mL	40 mL	2 x 80 mL	40 mL	2 x 40 mL
Buffer TN	20 mL	130 mL	55 mL	2 x 130 mL	20 mL	1 x 55 mL 1 x 20 mL
Wash Solution E	12 mL	2 x 18 mL	–	–	–	–
Elution Buffer K	8 mL	30 mL	–	–	8 mL	2 x 8 mL
Wash Solution J	–	–	25 mL	3 x 25 mL	–	–
Elution Buffer J	–	–	24 mL	120 mL	–	–
RNase A	1 vial	1 vial	1 vial	1 vial	1 vial	1 vial
Magnetic Bead Suspension	–	–	–	–	1 x 1.1 mL	4 x 1.1 mL
Spin Columns	50	250	–	–	–	–
Collection Tubes	50	250	–	–	–	–
DNA Maxi Spin Columns with Collection Tubes (Clear ring in column)	–	–	4	20	–	–
Maxi Spin Filter Columns with Collection Tubes (Grey ring in column)	–	–	4	20	–	–
96-Well Plate	–	–	–	–	–	2
Elution Tubes (1.7 mL)	50	250	–	–	50	–
Elution Tubes (50 mL)	–	–	4	20
96-Well Elution Plate	–	–	–	–	–	2
Adhesive Tape	–	–	–	–	–	2
Product Insert	1	1	1	1	1	1