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 V3-V4 Library Preparation Kit for Illumina consists of the reagents and components required for library preparation of the 16S V3-V4 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 V3-V4 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 V3-V4 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

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 V3-V4 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. 70400 (24 preps)	Cat. 70410, 70420, 70430, 70440 (96 preps)
Amplicon PCR (PCR 1)	MGX Master Mix	330 µL	1,320 µL
	16S V3-V4 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

Description 

The DM3100 ExcelBand™ 1 KB (0.25-10 kb) DNA Ladder is a ready-to-use DNA ladder, which is pre-mixed with loading dye for direct gel loading. The DNA ladder DM3100 is composed of 13 individual DNA fragments: 10k, 8k, 6k, 5k, 4k, 3k, 2.5k, 2k, 1.5k, 1k, 750, 500, and 250 bp derived from a mixture of PCR products and specifically digested plasmid DNA. This product contains two enhanced bands (3 kb and 1 kb) for easier reference. In addition, two tracking dyes, Xylene cyanol FF and Bromophenol blue which mimic the migration of 4,000 bp and 500 bp dsDNA during electrophoresis are also added for real time monitoring. 

Features

• Sharp bands
• Quick reference— enhanced bands 
• Ready-to-use— premixed with loading dye for direct loading
• Stable— room temperature storage over 6 months

Source 

Phenol extracted PCR products and dsDNA digested with specific restriction enzymes, equilibrated in 10 mM Tris-HCl (pH 8.0) and 10 mM EDTA. 

Range 

250 ~ 10,000 bp 

Concentration 

50 µg/ 500 µl 

Recommended loading volume 

5 µl/ well 

Storage

Room temperature for 6 months
4°C for 12 months 
-20°C for 36 months

The DM3100 ExcelBand™ 1 KB (0.25-10 kb) DNA Ladder is a ready-to-use DNA ladder, which is pre-mixed with loading dye for direct gel loading. The DNA ladder DM3100 is composed of 13 individual DNA fragments: 10k, 8k, 6k, 5k, 4k, 3k, 2.5k, 2k, 1.5k, 1k, 750, 500, and 250 bp derived from a mixture of PCR products and specifically digested plasmid DNA. This product contains two enhanced bands (3 kb and 1 kb) for easier reference. In addition, two tracking dyes, Xylene cyanol FF and Bromophenol blue which mimic the migration of 4,000 bp and 500 bp dsDNA during electrophoresis are also added for real time monitoring. 

Introduction

HiPure Insect DNA Kits provides a simple and rapid solution for total DNA extraction of insect tissue samples. This kit is based on silica gel column purification technology without toxic phenol chloroform extraction and time-consuming alcohol precipitation. The whole extraction process only takes 30 minutes. HiPure Insect DNA Kit can process tissue samples less than 10mg at a time. Hipure Insect DNA 96 kit can process 96 insect tissue samples at a high throughput. The obtained DNA can be directly used in PCR, Southern blot, viral DNA detection and other experiments.

Details

Specifications

Features	Specifications
Main Functions	Isolation total DNA from insect tissue
Applications	PCR, southern bolt and virus detection, etc
Purification method	96 well DNA plate
Purification technology	Silica technology
Process method	Manual (centrifugation or vacuum)
Sample type	Insect tissue samples
Sample amount
Elution volume
Time per run
Liquid carrying volume per column
Binding yield of column

Principles

This product is based on silica column purification. The sample is lysed and digested with lysate and protease, DNA is released into the lysate. Transfer to an adsorption column. Nucleic acid is adsorbed on the membrane, while protein is not adsorbed and is removed with filtration. 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 DNA – can be used in sensitive downstream applications such as multiplex and quantitative pcr
• Good repeatability – suitable for extracting high-yield DNA from insect tissue samples
• Safe – without phenol chloroform extraction and alcohol precipitation

Kit Contents

Contents	D313901	D313902
Purification Times	1 x 96	4 x 96
HiPure DNA Plate	1	4
2.2 ml Collection Plate	1	4
1.6 ml Collection Plate	1	4
0.5ml Collection Plate	1	4
Seal Film	8	32
Buffer ITL	30 ml	120 ml
Buffer IL	30 ml	125 ml
Buffer GW1	44 ml	2 x 110 ml
Buffer GW2	50 ml	3 x 50 ml
Proteinase K	50 mg	200 mg
Protease Dissolve Buffer	6 ml	15 ml
Buffer AE	20 ml	60 ml

Storage and Stability

Proteinase K, RNase A should be stored at 2-8°C upon arrival. However, short-term storage (up to 12 weeks) at room temperature (15-25°C) does not affect their performance. The remaining kit components can be stored at room temperature (15-25°C) and are stable for at least 18 months under these conditions. The entire kit can be stored at 2–8°C, but in thiscase buffers should be redissolved before use. Make sure that all buffers areat room temperature when used.

HiPure Insect DNA Kits provides a simple and rapid solution for total DNA extraction of insect tissue samples. This kit is based on silica gel column purification technology without toxic phenol chloroform extraction and time-consuming alcohol precipitation. The whole extraction process only takes 30 minutes. HiPure Insect DNA Kit can process tissue samples less than 10mg at a time. Hipure Insect DNA 96 kit can process 96 insect tissue samples at a high throughput. The obtained DNA can be directly used in PCR, Southern blot, viral DNA detection and other experiments.