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Presto™ FFPE RNA Mini Kit (RFD050, RFD100) with DNase

RNA Extraction/Total RNA

The Presto™ FFPE RNA Mini Kit was designed specifically for purifying total RNA from formalin-fixed, paraffin-embedded (FFPE) tissue. Paraffin is initially dissolved with organic solvents such as Xylene followed by Proteinase K digestion. RNA in chaotropic salt is then bound by the glass fiber matrix of the spin column and DNA contamination is removed using an in column DNase treatment. Other contaminants are removed using Wash Buffer and the purified total RNA is eluted in RNase-free Water. High quality total RNA can be purified within 80 minutes without phenol extraction or alcohol precipitation. The purified RNA is ready for use in Real-time PCR, mRNA selection, cDNA synthesis and NGS.

DOCUMENTS

COMPLETE PROTOCOL

DESCRIPTION

Specifications (Cat. # RFD050, RFD100)

Sample: up to 25 mg sections of FFPE, 4~10 sections of 5~20 μm thick FFPE
High Yield: up to 50 μg of total RNA
Spin Columns: glass fiber membrane optimized for total RNA extraction
Individually packaged RNA spin columns and collection tubes, certified RNase and DNase-free
Storage: Storage: dry at room temperature (15-25°C), DNase I is shipped at room temperature and should be stored at -20°C
Operation Time: within 80 minutes

APPLICATIONS

Real-time PCR, mRNA Selection, cDNA Synthesis, NGS

COMPONENTS

RT Buffer
RB Buffer
Proteinase K
DNase I
DNase I Reaction Buffer
W1 Buffer
Wash Buffer
RNase-free Water
RB Columns
2 ml Collection Tubes

QUALITY CONTROL

The quality of the Presto™ FFPE RNA Mini Kit is tested on a lot-to-lot basis according to Geneaid's ISO-certified quality management system. Total RNA is isolated from 25 mg sections of FFPE, quantified with a spectrophotometer and analyzed by electrophoresis.

Presto^TM FFPE RNA Mini Kit Test Data

Figure 1 .(A)Total RNA was isolated from 4 slices of chicken FFPE kidney section (5 μm thickness) using Geneaid PrestoTM FFPE RNA Mini Kit. The RNA integrity was analyzed by capillary electrophoresis (BIOptic, Qsep100). The RQN (RNA Quality Number) = 7.4 indicated the intact RNA was purified. However, due to effects of formalin fixation, embedding and storage, RNA from FFPE samples will usually show varying degrees of degradation. Typical RQN of RNA purified with PrestoTM FFPE RNA Mini Kit are in range of 2–6. (B)Since the RIN values from degraded FFPE samples are not a sensitive measure of RNA quality. Instead, the DV200 (percentage of RNA fragments that are greater than 200 nucleotides) developed by Illumina is used to assess FFPE RNA quality. In this case, the DV200 of purified RNA is 90.4%. The data showed that more than 90% of large RNA fragments in the purified RNA are excellent for successful library preparation. In general, the DV200 of purified RNA are in range of 30%-90% using Geneaid PrestoTM FFPE RNA Mini Kit.

Figure 2 .High efficiency in real-time PCR.Total RNA was extracted from 2-5 slices of 5-20 μm thick FFPE sections from 15 normal endometrium patients using Geneaid PrestoTM FFPE RNA Mini Kit. The TET1 and GAPDH mRNA expression level were measured by real time one-step RT-PCR. The data showed that the purified RNA with excellent performance in downstream application such as RT-PCR.

Total RNA Extraction Kits Specifications

Specifications	Total RNA Mini Kit Blood Cultured Cell	Total RNA Mini Kit Tissue	Total RNA Mini Kit Plant	Presto™ Mini RNA Bacteria Kit	Presto™ Mini RNA Yeast Kit	miRNA Isolation Kit	GENEzol™ TriRNA Pure Kit
Cat. #	RBD050 RBD100 RBD300	RTD050 RTD100 RTD300	RPD050 RPD100 RPD300	RBBD050 RBBD100 RBBD300	RBYD050 RBYD100 RBYD300	RMI050	GZXD050 GZXD100 GZXD200
Format	Spin Column	Spin Column	Spin Column	Spin Column	Spin Column	Spin Column	Tri Reagent Spin Column
Binding Capacity	50 µg	50 µg	50 µg	50 µg	50 µg	50 µg	50 µg
Sample	blood, cells	animal tissue	plant tissue	bacterial cells	yeast cells	animal tissue, cells	blood,cells, tissue, body fluids, bacteria
Micropestle	N/A	Yes	N/A	N/A	N/A	Yes	N/A
Filter Column	N/A	Yes	Yes	N/A	N/A	N/A	N/A
Enzyme	Dnase I	Dnase I	Dnase I	Lysozyme Dnase I	Dnase I	N/A	Dnase I
Typical Yield	2-3 µg from 300 µl blood, 25-30 µg from 10⁶ cells	5-30 µg	5-30 µg	40-50 µg from 1x10⁹ bacterial cells	15-20 µg from 5x10⁷ yeast cells	5-30 µg	5-30 µg
Elution Volume	50 µl	50 µl	50 µl	50 µl	50 µl	50 µl	25-50 µl
Time	<20 min.	<15 min.	<20 min.	<20 min.	<20 min.	<30 min.	<15 min.

RELATED PRODUCTS

Product Name	RXNS	Catalogue Number
Total RNA Mini Kit (Blood/Cultured Cell)	50/100/300	RB050/100/300, RBD050/100/300
Total RNA Mini Kit (Tissue)	50/100/300	RT050/100/300, RTD050/100/300
Total RNA Mini Kit (Plant)	50/100/300	RP050/100/300, RPD050/100/300
Presto™ Mini RNA Bacteria Kit	50/100/300	RBB050/100/300, RBBD050/100/300
Presto™ Mini RNA Yeast Kit	50/100/300	RBY050/100/300, RBYD050/100/300
miRNA Isolation Kit	50	RMI050
GENEzol™ TriRNA Pure Kit	50/100/200	GZX050/100/200, GZXD050/100/200
GENEzol™ 96 Well TriRNA Pure Kit	4 x 96/10 x 96	96GZX04/10

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