DBCO-NHCO-PEG4-acid

Overview

• Isolate all sizes of circulating and exosomal RNA, including microRNA
• Versatile urine input ranges
• No phenol extractions
• No carrier RNA
• Bind and elute all RNA irrespective of size or GC content, without bias
• Concentrate circulating RNA and exosomal RNA into flexible elution volumes
• Purified RNA is suitable for a variety of downstream applications, including Small RNA Sequencing. Find out more information on Norgen’s NGS services
• Purification is based on spin column chromatography that uses Norgen’s proprietary resin separation matrix

These kits provide a fast, reliable and convenient method to purify and concentrate high quality, high purity and inhibitor-free cell-free circulating RNA, including exosomal RNA as well as viral RNA from fresh, preserved or frozen urine samples. All components for the purification are provided in one convenient and fast kit for the easy processing of small input volumes of bodily fluids. The purified urine RNA is fully compatible with all downstream applications including PCR, qPCR, methylation-sensitive reverse transcription qPCR, reverse transcription PCR, Northern blotting, RNase protection and primer extension, expression array assays, and NGS.

Background

Recent evidence indicates that cell-free circulating RNA (cf-RNA) including exosomal RNA in urine contains valuable information for the discovery of biomarkers that can help for the early detection of certain cancer types and for monitoring the disease status as well as for the detection of any infectious pathogens. Exosomes can be found in saliva, blood, urine, amniotic fluid and malignant ascitic fluids, among other biological fluids. Evidence has been accumulating recently that these vesicles act as cellular messengers, conveying information to distant cells and tissues within the body. These exosomes may play a functional role in mediating adaptive immune responses to infectious agents and tumours, tissue repair, neural communication and transfer of pathogenic proteins. For this reason exosomal RNAs may serve as biomarkers for various diseases including cancer. The advantage for using urine as a source for cancer biomarkers is that it can be acquired in large quantities without using invasive procedures. In addition, repeated sampling from the same individual is applicable, which facilitates longitudinal studies. There are many advantages favouring the use of urinary nucleic acid for cancer biomarker discovery over blood, tissue samples or other bodily fluids, including: (1) urine is non-infectious for HIV and less infectious for many other pathogens; (2) the profile of urinary nucleic acid is similar to that in plasma or serum but with a lower concentration; (3) Nucleic acid purification from urine is technically much easier because of its low protein concentration (1000-fold lower than blood).

Urine Cell-Free Circulating RNA Purification Mini Kit

For sample volumes ranging from 250 µL to 2 mL.

Urine Cell-Free Circulating RNA Purification Midi Kit

For sample volumes ranging from 2 mL to 10 mL. The first column will handle the large volume input of urine that is followed by a concentration on a mini column for a final elution of 50 µL to 100 µL.

Urine Cell-Free Circulating RNA Purification Maxi Kit

For sample volumes ranging from 10 mL to 30 mL. The first column will handle the large volume input of urine that is followed by a concentration on a mini column for a final elution of 50 µL to 100 µL

Details

Supporting Data

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Kit Specifications
Minimum Urine Input	10 mL
Maximum Urine Input	30 mL
Size of RNA Purified	All sizes including miRNA and small RNA (< 200 nt)
Elution Volume	50-100 μL
Time to Complete 10 Purifications	40-45 minutes
Average Yields	Variable depending on specimen

*Please check page 6 of the product insert for average yields and common RNA quantification methods.

Storage Conditions and Product Stability
All buffers should be kept tightly sealed and stored at room temperature. These kits are stable for 2 years after the date of shipment. It is recommended to warm Lysis Buffer A for 20 minutes at 60°C if any salt precipitation is observed.

Component	Cat. 56900 (50 preps)	Cat. 57000 (20 preps)	Cat. 57100 (10 preps)
Binding Solution K	25 mL	75 mL	1 x 75 mL 1 x 25 mL
Lysis Buffer A	30 mL	20 mL	20 mL
Wash Solution A	18 mL	18 mL	18 mL
Elution Solution A	6 mL	6 mL	6 mL
Mini Spin Columns	50	20	10
Midi Spin Columns	–	20	–
Maxi Spin Columns	–	–	10
Collection Tubes	50	20	10
Elution Tubes (1.7 mL)	50	20	10
Product Insert	1	1	1

The Opentrons 24-well Aluminum Block can be placed directly on the OT-2/Opentrons Flex deck or on the Opentrons Temperature Module. This block acts as a 24 position rack to keep 2mL tubes, 1.5mL tubes, and the NEST 0.5mL tubes at constant temperature. The aluminum block holds temperatures between 4°C and 95°C when used with the Opentrons Temperature Module (GEN2).

The Opentrons 24-well Aluminum Block can be placed directly on the OT-2/Opentrons Flex deck or on the Opentrons Temperature Module. This block acts as a 24 position rack to keep 2mL tubes, 1.5mL tubes, and the NEST 0.5mL tubes at constant temperature. The aluminum block holds temperatures between 4°C and 95°C when used with the Opentrons Temperature Module (GEN2).

The kit was developed for construction of high quality libraries for next generation sequencing (illumina and MGI Platforms). The kit needs double strand DNA fragments (blunt and/or sticky) as input DNA for NGS library preparation, and is compatible with DNA fragments generated from both enzymatic methods and physical methods (sonication, nebulization etc.). Library multiplexing is possible with different types of indexes.

The kit was optimized for next generation sequencing (NGS) library preparation with different types of samples. Most of DNA library preparation requires the ligation of sheared DNA fragments to library adaptors and the DNA library preparation is closely related to the quality of NGS data. With BioDynami’s unique DNA library preparation technologies, the fast and simple kit allows high quality NGS library preparation to be completed in 1.5 hours with only 10 minutes of hands-on time.

<img decoding="async" src="https://biodynami.com/wp-content/uploads/2020/07/NGS-DNA-Library-Prep-Kit-illumina-1.png" alt=""

Some genomic regions are very difficult to be covered evenly and usually result in very low coverage rate or gap in these regions.

Typical difficult regions are:
• with high GC contents
• have secondary structures: mainly due to repeat sequences
• the worst cases: have both high GC contents and repeated sequences.

Example: human TERT gene is one of the most difficult regions as shown above. NGS data showed that BioDynami kit has the best performance to cover the extremely difficult human TERT gene region.

Limited GC bias across whole genome

<img decoding="async" src="https://biodynami.com/wp-content/uploads/2020/07/BioDynami-NGS-DNA-Library-Prep-Kit-GC-bias.png" alt="

Three index types are available for the illumina platform kits:

Non-index (illumina Cat.# 30009): Libraries do not have index.

Index (illumina Cat.# 30021): A unique barcode sequence with 6 bases has been included in each of the index primers. RNA Sequencing library multiplexing is possible with up to 48 samples. Index information can be downloaded here.

Unique dual index (illumina Cat.# 30023): RNA Sequencing library multiplexing up to 96 samples is possible with the unique dual indexes. We have developed a 4-Base Difference Index System. The system can generate indexes with at least 4 bases different from others in the 8-base indexing region. the unique dual indexing primers identify sequencing errors such as index hopping, mis-assignment, and de-multiplexing errors. Index information can be downloaded here.

Indexes are available for the MGI platform kits (Cat.# 34021).

The kit was developed for construction of high quality libraries for next generation sequencing (illumina and MGI Platforms). The kit needs double strand DNA fragments (blunt and/or sticky) as input DNA for NGS library preparation, and is compatible with DNA fragments generated from both enzymatic methods and physical methods (sonication, nebulization etc.). Library multiplexing is possible with different types of indexes.