Plant/Fungi RNA Purification Kit

Supplier
Norgen Biotek
Cat. no.
1025800
Kit size
50
Links
Norgen Biotek Protocol Information sheet

Features

• No liquid nitrogen required for homogenization - Liquid nitrogen is not required for homogenization of samples, making RNA purification rapid and convenient.
• Isolate RNA from a wide range of samples - Total RNA can be isolated from a wide range of plant and filamentous fungi samples.
• Isolate a diversity of RNA species - All sizes of RNA are isolated from large mRNA down to microRNA, without the use of phenol or chloroform.
• Isolate total RNA, including viral RNA - RNA samples can be used for the detection of viral pathogens, as viral RNA is isolated with the total RNA.
• High yield of RNA - High yields of purified RNA can be isolated with this kit.
• No phenol:chloroform extractions - Total RNA is isolated without the use of harmful chemicals such as phenol or chloroform. RNA is of the highest quality and can be used in a number of downstream applications.

Norgen's Plant/Fungi Total RNA Purification Kit provides a rapid method for the isolation and purification of total RNA, including viral RNA, from a wide range of plant and filamentous fungal species. Total RNA can be purified from fresh or frozen plant tissues, plant cells or filamentous fungi samples using this kit. All sizes of RNA are purified, from large mRNA and ribosomal RNA down to microRNA (miRNA) and small interfering RNA (siRNA). The procedure is rapid and convenient, as it does not rely on the use of liquid nitrogen in order to homogenize the samples. Purification is based on spin column chromatography using Norgen’s proprietary resin as the separation matrix. The RNA is preferentially purified from other cellular components, such as proteins, without the use of phenol or chloroform. The purified RNA is of the highest integrity, and can be used in a number of downstream applications including real time PCR, reverse transcription PCR, Northern blotting, RNase protection and primer extension, and expression array assays.

Purification is based on spin column chromatography using Norgen’s proprietary resin as the separation matrix. The RNA is preferentially purified from other cellular components such as proteins without the use of phenol or chloroform. The process involves first macerating the cells or tissue in a mortar with the provided Lysis Solution (please see the flow chart on page 4). The Lysis Solution contains detergents, as well as large amounts of a chaotropic denaturant that will rapidly inactivate RNases and proteases that are present. Alternatively, liquid nitrogen can be used to homogenize the sample. The lysate is then spun in a microcentrifuge in order to pellet and remove any debris. Ethanol is then added to the clarified lysate, and the solution is loaded onto a spin-column. Norgen’s resin binds nucleic acids in a manner that depends on ionic concentrations, thus only the RNA will bind to the column while most of the DNA and proteins are removed in the flowthrough. The bound RNA is then washed with the provided Wash Solution in order to remove any remaining impurities, and the purified total RNA is eluted with the Elution Buffer. The purified RNA is of the highest integrity, and can be used in a number of downstream applications.

Column Binding Capacity	50 µg
Maximum Column Loading Volume	600 µl
Size of RNA purified	All sizes, including small RNA (<200 nt)
Maximum Amount of Starting Material:	Plant Tissues: 50 mg	Plant Cells: 1x10exp6 cells	Fungi: 50 mg (wet weight)
Average Yields*	50 mg Tomato Leaves: 60 µg	50 mg Tobacco Leaves: 60 µg	50 mg Plum Leaves: 32 µg	50 mg Grape Leaves: 35 µg	50 mg Peach Leaves: 30 µg	* average yields will vary depending upon a number of factors including species, growth conditions used and developmental stage.
Time to Complete 10 Purifications	30 minutes

Isolate RNA from a Wide Range of Plants

RNA was isolated from 50 mg samples of tobacco leaves (Lanes 1 and 2), tomato leaves (Lanes 3 and 4), peach leaves (Lanes 5 and 6) and grape leaves (Lanes 7 and 8) using Norgen’s Plant/Fungi Total RNA Purification Kit, and 5 uL aliquots were run on a 1X MOPS 1 % formaldehyde-agarose gel. Total RNA was isolated in all cases, including all small microRNA species Lane M is Norgen’s 1 kb RNA Ladder.

Detection of Plum Pox Virus in Peach Leaves

Total RNA was isolated from two 50 mg samples of peach leaves using Norgen’s Plant/Fungi Total RNA Purification Kit, and 3 m L of the RNA was used in an RT-PCR reaction for the detection of plum pox virus. Plum pox virus was detected from both samples, indicating that the RNA is of a high quality and that the kit is highly sensitive for total RNA isolation.

• Quantitative, real-time RT-PCR
• RT-PCR
• Northern blotting
• RNase protection and primer extension
• Expression array assays