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-V3 Library Preparation Kit for Illumina consists of the reagents and components required for library preparation of the 16S V1-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 V1-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 V1-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 V1-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. 70200 (24 preps)	Cat. 70210, 70220, 70230, 70240 (96 preps)
Amplicon PCR (PCR 1)	MGX Master Mix	330 µL	1,320 µL
	16S V1-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

This product is specially designed for DNA extraction from forensic samples such as bone, hair, fingernail, seminal strain, dry blood spot, ect. Especially designed for forensic detection. The obtained DNA can be directly used for PCR, STR detection and other down stream applications.

Details

Kit Contents

Contents	D635901D	D635902D	D635903D
Purification Times	48	96	480
MagPure Particles N	1.1 ml	2.2 ml	11 ml
Buffer ATL	40 ml	80 ml	2 x 200 ml
Buffer BGL	40 ml	80 ml	2 x 200 ml
Buffer BST1	60 ml	120 ml	550 ml
Buffer GW2*	20 ml	25 ml	2 x 50 ml
Proteinase K	48 mg	110 mg	2 x 220 mg
Protease Dissolve Buffer	5 ml	10 ml	30 ml
DTT	235 mg	235 mg	3 x 235 mg
Elution Buffer	10 ml	10 ml	30 ml

Prefilled Kit Contents

32/48 channel machine

Product	Contents and volume	D6359D-TL-06
Buffer BGL		80 ml
Buffer ATL		80 ml
Proteinase K		110 mg
Protease Dissolve Buffer		10 ml
DTT Powder		2 x 235 mg
Elution Buffer		5 ml
AS-Tip		12 pcs
2.0ml V bottom plate	Row 1/7：empty	6 plates
	Row 2/8：500μl Buffer BST1
	Row 3/9：500μl Buffer BST1
	Row 4/10：20μl MagPure Particles N 500μl Buffer GW2 Row 5/11：500μl Buffer GW2 (ethanol ~350ul)
	Row 6/12：60μl Elution Buffer

96 channel machine

Product	Contents and volume	D6359D-F-96	D6359D-F-48
Buffer BGL		80 ml	40 ml
Buffer ATL		80 ml	40 ml
Proteinase K		110 mg	48 mg
Protease Dissolve Buffer		10 ml	5 ml
DTT Powder		235 mg	235 mg
Elution Buffer		5 ml	5 ml
96-Tip (AS)		1	1
Sample Plate 1	500μl Buffer BST1	1	1
Sample Plate 2	500μl Buffer BST1	1	1
Wash Plate 1	20μl MagPure Particles N500μl Buffer GW2	1	1
Wash Plate 2	500μl Buffer GW2	1	1
Elute Plate	60μl Elution Buffer	1	1

Storage and Stability

Proteinase K and DTT Powder 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 condition.

This product is specially designed for DNA extraction from forensic samples such as bone, hair, fingernail, seminal strain, dry blood spot, ect. Especially designed for forensic detection. The obtained DNA can be directly used for PCR, STR detection and other down stream applications.

DBCO-PEG4-NHS Ester is a click chemistry PEG reagent containing NHS ester that is able to react specifically and efficiently with primary amines (e.g. the side chain of lysine residues or aminosilane-coated surfaces) at neutral or slightly basic condition to form a covalent bond. The hydrophilic PEG spacer arm improves water solubility and provides a long and flexible connection that minimizes steric hindrance involved with ligation. DBCO is commonly used for copper-free Click Chemistry reactions. Reagent grade, for research purpose. Please contact us for GMP-grade inquiries.

DBCO-PEG4-NHS Ester is a click chemistry PEG reagent containing NHS ester that is able to react specifically and efficiently with primary amines (e.g. the side chain of lysine residues or aminosilane-coated surfaces) at neutral or slightly basic condition to form a covalent bond. The hydrophilic PEG spacer arm improves water solubility and provides a long and flexible connection that minimizes steric hindrance involved with ligation. DBCO is commonly used for copper-free Click Chemistry reactions. Reagent grade, for research purpose. Please contact us for GMP-grade inquiries.