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GenomONE - Neo EX Unique, efficient transfection of siRNA, DNA, oligonucleotides, proteins.Low toxicity with primary cells. Protocols for in vivo transfection.

GenomONE™ - Neo EX HVJ Envelope Transfection Kit

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Background

HVJ (hemagglutinating virus of Japan) Envelope VECTOR KIT is a tool for transfection of molecules (plasmid DNAs, siRNAs, oligonucleotides, proteins, antibodies etc.) into cells and animal tissue  by means of membrane fusion.
The HVJ envelope, carrying the molecule to be transfected, is composed of a completely inactivated and purified HVJ (Sendai virus) while preserving the cell membrane-fusing capability of the envelope.
GenomONE-Neo EX provides ready-to-use kits containing the HVJ envelope and auxiliary reagents (incorporation enhancer, incorporation reagent, introduction enhancer, buffer).

GenomONE products [Transfection, Cell fusion] Flyer
GenomONE products
[Transfection, Cell fusion] Flyer
GenomONE-Neo EX is novel and unique transfection kit which employ the membrane fusion ability of the envelope of Sendai virus (Hemagglutinating virus of Japan: HVJ). HVJ Envelope (HVJ-E) is a nonproliferative and noninfectious vesicle purified after complete inactivation of Sendai virus genomic RNA. HVJ-E can introduce a variety of molecules (plasmid DNA, siRNA/miRNA, protein) into cells via membrane fusion.

Principle

What is HVJ Envelope (HVJ-E)?
HVJ Envelope is a purified product prepared through complete inactivation of Sendai virus (HVJ*). It is a vesicle in which only the cell membrane-fusing capability of the envelope protein of Sendai virus is retained.

It is known that the HN protein in the tunica externa of the Sendai virus recognizes the receptor (possessing sialic acid at the terminal of sugar chain) on the cell membrane and adsorbs it, leading to the induction of membrane fusion mediated by F protein (another component of the envelope). The genomic RNA of the Sendai virus contained in HVJ-E has been inactivated completely and has neither infective nor proliferative potentials in humans or experimental animals. HVJ-E can be used safely at ordinary laboratories, without requiring any special operations or facilities.
*HVJ:Hemagglutinating Virus of Japan.

  • HVJ-E can incorporate various molecules (plasmid DNA, siRNA/miRNA, proteins)
  • HVJ-E-incorporated molecules are introduced into cells via membrane fusion
  • HVJ-E is also applicable to in vivo transfection

Conventional non-viral transfection tools, including cationic lipids, are incorporated into cells through endocytosis which results in degradation of the transferred DNA by lysosomes.
On the other hand, HVJ Envelope VECTOR resists degradation by lysosomes, making it easy to transfer the specified DNA. Therefore, HVJ Envelope VECTOR yields highly efficient gene expression.
Sialic acid receptors, which are needed to trigger binding with HN protein, exist in almost all animal cells. Thus, HVJ Envelope VECTOR is useful for a wide range of targets.

Features and Advantages

GenomONE-Neo EX
unique, exciting transfection kit for cells, tissue, and live animals.

Use GenomONE-Neo EX: for:

siRNA
strong knockdowns with low pM amounts of siRNA. Chemical modification of siRNA not required; means high functional expression..

DNA
small and large, linear or circular

Oligonucleotides
DNA, RNA

Proteins
enzymes and antibodies retain function and specificity

In vivo transfection!!
protocol and literature citations for effectiveness of molecules transfected into live animals following direct injection of HVJ-E vector.

High throughput screening
transfect multiple molecules simultaneously

 

1. Wide usability
GenomONE-Neo EX HVJ Envelope VECTOR KIT is a highly flexible tool for transfecting a wide variety of molecules (plasmid DNAs, siRNAs, oligonucleotides, proteins, antibodies etc.) into cultured cells (adherent and non-adherent) and tissue. GenomONE-Neo EX is useful for transfecting sensitive primary cells and is further distinguished by its ability to deliver contents into live animals. Many literature citations are available for each GenomONE-Neo EX application. 

2. Safety
Unlike cationic lipids, GenomONE-Neo EX delivers the molecule of interest into cells via membrane fusion, not by endocytosis where cargo may be degraded by lysosomal enzymes. Since GenomeONE-Neo is a purified HVJ (Sendai virus) product, prepared from virus particles completely inactivated for infectious ability and proliferative potential it is completely safe to use without special precautions. 

3. Easy
GenomONE-Neo EX provides ready-to-use kits containing the HVJ envelope and required auxiliary reagents (incorporation enhancer, incorporation reagent, introduction enhancer, and buffer). Suggested protocols for all major applications are included.

Comparison data of GenomONE-Neo with cationic lipid-based reagents (liposome)

1. Comparison of GenomONE-Neo with cationic lipid-based reagent for siRNA delivery

1. Phospholamban (PLB) gene knockdown in primary rat myocardial cell cultures

GenomONE-Neo

1: Non-treated
2: PLB siRNA (10µg )
3: PLB siRNA (2µg)
4: Scramble siRNA (10µg)
5: HVJ-E only

Cationic lipid reagent

1: PLB siRNA (10µg )
2: PLB siRNA (2µg)
3: Scramble siRNA (10µg )
4: Non-treated

The efficiency of siRNA transfer into cells was high (80-100%) with both reagents.  However, compared to cationic lipid, GenomONE-Neo  induced knock-down of the target protein more efficiently at lower concentrations of siRNA.

⟨Data⟩ Drs. M. Arai and A. Watanabe (Gunma University Graduate School of Medicine)
⟨Related article⟩ Watanabe et al., J. Mol. Cell. Cardiol., 37 (3), 691-698 (2004)

2 Bin1 gene knockdown in C2C12 (mouse myoblast cell line) after induction of differentiation

Use of conventional representative siRNA transfection reagents (products A, B, and C) failed to exert sufficient knock-down effects, demonstrating the superiority of GenomONE-Neo to these products.

⟨Data⟩ Drs. C. Kojima* and H. Sabe (Osaka Bioscience Institute, *Current address: Osaka Prefecture University)
⟨Related article⟩ Kojima et al.,  EMBO J., 23, 4413-4422 (2004).

3 Eg5 gene knockdown in various cell cultures

Eg5-siRNA was incorporated in each reagent to a final concentration of 50 nM in a 96-well plate. Cells of various lines were treated with the reagents. Forty-eight hours later, the percentage of viable cells was measured by WST-1 assay. Eg5 knockdown resulted in suppression of cell proliferation and induced apoptosis. The finding of a lower percentage of viable cells indicated stronger Eg5 knockdown effects.

4 EGFP gene knockdown in monkey ES cells
Biochem. Biophys. Res. Commun., 331, 1039-1044 (2005).

2. Highly efficient in vivo gene transfer to the mouse uterus
-Comparison data with cationic lipid-based reagent-
Mol. Hum. Reprod., 9, 603-609 (2003)

3.Transfection of neuroprogenitor cells with iron nanoparticles for magnetic resonance imaging tracking
-Comparison data with cationic lipid-based reagent-
Mol. Imaging Biol., 7, 286-295 (2005).


Application

in vivo

Title Target cell/Tissues Introduced molecule
Intrathecal administration of siRNA to rats Rat lumbar dorsal horn siRNA
Retrograde Injection of siRNA into Rat Submandibular Gland (in vivo) Rat submandibular gland

Excerpt from "Intrathecal administration of siRNA to rats"

By using HVJ-E vector, NK1R siRNA was administered into the spinal cord. Immunohistochemical reaction of NK1R was suppressed by siRNA-treated during 1 to 7 days.

[Data]
  • Dr. Rumi Naono-Nakayama
    Division of Anatomy and Cell Biology, Faculty of Medicine, Tohoku Medical and Pharmaceutical University (Japan)
  • Dr. Toshikazu Nishimori
    Department of Psychiatry, Faculty of Medicine, University of Miyazaki (Japan)
[Related article]
  • Eur. J. Pharmacol., 670, 448-457 (2011).

in vitro

Title Target cell/Tissues Introduced molecule
Phospholamban (PLB) knock-down in primary rat myocardial cell cultures Neonatal rat myocardial cells (primary cultures) siRNA
RNAi in difficult-to-transfect U937 cells U937

Intracellular delivery of Cre recombinase (protein)

By introducing Cre recombinase (protein) into 2-2 cells with GenomONE-Neo, loxp sites inserted in the genome sequence were deleted and Lac Z gene expression was induced.
2-2 (monkey, African green, RIKEN BioResource Center): 35 copies of PCAG-loxP-neopA-loxP-LacZ were tandemly inserted into the genome.

TOPICS (Representative Application Examples)
Applications for difficult-to-transfect cells and primary cells as well as in vivo experiments.

GenomONE-NEO EX siRNA-data
GenomONE-NEO EX siRNA-data
GenomONE-NEO EX Protein-data
FAQ GenomONE-NEO EX [FD]
GenomONE-NEO EX Protein-data
High-throughput screening Data sheet
GenomONE-NEO EX Protein-data
Reference list (in vitro)
GenomONE-NEO EX Protein-data
Reference list (in vivo)

 

[1] siRNA transfection

in vitro
Primary T cell (Human peripheral blood)
Primary cardiac myocyte (Rat cardiac myocyte)
Differentiated C2C12 (Mouse myoblast)
monkey ES cells
Min6 (Mouse pancreatic β cell)
U937 (Human myelomonocytic cell)

in vivo
ntradermally transplanted human cervix cancer /HeLa (SCID mouse)
Submandibular gland (rat)
 

[2] Protein delivery

in vitro
Primary macrophage (C3H mouse peritoneal resident)
Swiss 3T3 cell (Mouse embryonic fibroblasts)
 
in vivo
Nucleus tractus solitarius (Rat brain)
 

[3] Oligo DNA transfection

in vitro
Primary HDMECs (Human dermal microvascular endothelial cell)
Primary CD34+ cell (Human blood)
in vivo
Uterus (Mouse)
Skin, ear lobe (Mouse)
Lung (Mouse)
 

[4] Plasmid DNA transfection

in vivo
Uterus(Mouse)
Palatal periodontal tissue (Rat)
Myocardium (Rat heart)
Lung (neonatal porcine)
Subcutaneously transplanted human colon cancer /LoVo (nude mouse)
 

Protocol


GenomONE - Neo EX Instruction Manual

Kit component

GenomONE-Neo EX Kit Contents GN001EX GN004EX
Freeze-dried HVJ-E
(when reconstituted) 
0.26 ml 1 tube 4 tubes
reagent A 0.5 ml 1 tube 1 tube
reagent B 0.3 ml 1 tube 1 tube
reagent C 1.0 ml 1 tube 1 tube
Buffer 6.5ml 1 tube 1 tube

The HVJ Envelope VECTOR KIT "GenomONE-Neo EX" series consists of HVJ Envelope and sub reagent set. Sub reagent set consists of 3 reagents (reagent A, reagent B and reagent C) and buffer for suspension or dilution.

  • Since the HVJ envelope of GenomONE-Neo EX (HVJ-E(N)) has a high fusing capability, it allows completion of transfection through a shorter contact duration with cells or tissues and has more suitable characteristics as follows: the reduction of toxicity to sensitive primary cultured cells, and the use in vivo where the transfected molecule is likely to diffuse due to inter-tissue transfer.
  • Some cultured cell lines show intense cell-to-cell fusion when GenomONE-Neo EX is used.
  • Applicability of  GenomONE-Neo EX may vary depending on the type of cell and the features of the experimental system.

Technical Support

Product List

Product Name Cat# Quantity Price

GenomONE-Neo EX HVJ-E 1 vial Transfection Reagents

ISK-GN-001-EX 1SET Contact Us

GenomONE-Neo EX HVJ-E 4 vials Transfection Reagents

ISK-GN-004-EX 1SET Contact Us

To be used for research only. DO NOT use for human gene therapy or clinical diagnosis.

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