mNeonGreen | Fluorescent Protein
AS21 4525 | Clonality: Polyclonal | Host: Rabbit | Reactivity: mNeonGreen | Fluorescent Protein

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Product Information
Reactivity
Application examples

Samples:
1- Precision Plus Protein Standards (Bio-Rad)
2-Arabidopsis thaliana expressing mNeonGreen-tagged fusion protein
3-Arabidopsis thaliana wildtype total cell extract
4-Chlamydomonas reinhardtii expressing mNeonGreen-tagged fusion protein
5-Chlamydomonas reinhardtii wildtype cell extract
20 µg of total protein extracted from Arabidopsis thaliana leaves or Chlamydomonas reinhardtii cells with 62.5 mM Tris-HCl pH 6.8, 10 % glycerol, 2 % SDS, 50 mM DTT, 1x protease inhibitors (P9599, Sigma) and heated at 65ºC for 5 minutes were separated on 10 % SDS-PAGE and blotted 1 h to PVDF membrane (pore size 0.45 µm) using wet transfer. The blot was blocked with 5 % milk/TBS-T for 1 h/RT with agitation and then incubated in the primary antibody at a dilution 1: 1000 in 5 % milk/TBS-T for 1h/RT with agitation. The antibody solution was decanted, and the blot was briefly rinsed, then washed three times for 10 minutes in TBS-T with agitation. The blot was incubated in Agrisera matching secondary antibody (anti-rabbit IgG, HRP conjugated, AS09 602) at a dilution of 1: 25 000 in 5 % milk/TBS-T for 1h/RT with agitation. The blot was washed as above, developed for 2 minutes with AgriseraECLSuperBright substrate and imaged with a Bio-Rad ChemiDoc MP digital imaging system. Exposure time was 2.5 s.
Courtesy of Hanie Khorshidi, University of Saskatchewan, Canada
Samples:
1 - Nicotiana benthamiana whole leaf extract expressing mNeonGreen-ATG8
2 – Nicotiana benthamiana whole leaf extract expressing YFP-ATG8
3 – Nicotiana benthamiana whole leaf extract expressing mCherry-ATG8
MW markers: (Biorad)Precision Plus Protein Dual Color Standards, 500 µl #1610374
50 µg of total protein extracted freshly from Nicotiana benthamiana leaves with exaction buffer (50mM Tris-HCl pH7.5, 150mM NaCl, 10%(v/v) glycerol, 0.5mM EDTA and 0.5% NP40) denatured with SDS-sample buffer at 95°C for 5 min, proteins were separated on 10 % SDS-PAGE and blotted 0.5h to PVDF membrane using semi-dry transfer. Blots were blocked with 5 % milk in PBS-T for 1h/RT with agitation. Blot was incubated in the primary antibody at a dilution of 1: 3 000 for ON/4°C with agitation. The antibody solution was decanted, and the blot was rinsed briefly twice, then washed once for 15 min and 3 times for 5 min in PBS-T at RT with agitation. Blot was incubated in Agrisera matching secondary antibody (anti-rabbit IgG (H&L), horse radish peroxidase conjugated, AS09 602) diluted to 1: 5 000 in for 1h/RT with agitation. The blot was washed as above, developed for 5 mins with Agrisera ECL Bright and detected with film.
Courtesy of Dr. Jiaqi Sun, McGill University, Canada
Samples:
1- PageRuler™ Prestained Protein Ladder (Thermo Fisher)
2- 20 µg Arabidopsis thaliana wildtype microsomal membrane preparation
3- 20 µg Arabidopsis thaliana expressing VHA-a1-GSL-NeonGreen microsomal membrane preparation
4- 20 µg Arabidopsis thaliana expressing VHA-a3-GSL-NeonGreen microsomal membrane preparation
20 µg of microsomal membrane proteins were prepared from Arabidopsis thaliana 4-week -old rosette leaves with a buffer containing 350mM sucrose, 70mM Tris-HCl pH8, 10% glycerol, 3mM Na2EDTA, 0.15% BSA, 1.5% PVP-40, 4mM DTT, 1x Roche completeTM protease Inhibitor and denatured at 95ºC for 5 minutes. Proteins were separated on 10 % SDS-PAGE and blotted for 1 h to PVDF membrane (pore size 0.2 µm) using wet transfer. The blot was blocked with 5% milk/TBS-T (0.05%) for 1 h at RT with agitation and then incubated in the primary antibody at a dilution 1: 1000 in 5% milk/TBS-T (0.05%) for 1h at RT with agitation. The antibody solution was decanted, and the blot was briefly rinsed, then washed three times for 10 minutes in TBS-T (0.05%) with agitation. The blot was incubated in Agrisera matching secondary antibody (anti-rabbit IgG, HRP conjugated, AS09 602) at a dilution of 1: 25 000 in 5 % milk/TBS-T (0.05%) for 1hr at RT with agitation. The blot was washed as above, developed for 1 min with AgriseraECLBright substrate and imaged with a cooled CCD camera system (Intas ADVANCED Fluoreszenz u. ECL Imager). Exposure time was 10s.
Courtesy of Upendo Lupanga, Centre for Organismal Studies, University of Heidelberg, Germany
Additional information
- dCas9-P2A-DHFR(TSc3)-T2A-mNeonGreen [Cloning vector pBM020]
- Cas9-P2A-DHFR(TSc3)-T2A-mNeonGreen [Cloning vector pBM006]
- mNeonGreen4-tDeg [Cloning vector pminiCMV-mNeonGreen4-tDeg]
- ER-localized mNEONGREEN [Binary vector pKT-NM-erNEON]
- mNeonGreen-3xFLAG [Cloning vector pLM160-mNeonGreen]
- mNeonGreen-ty1 [Cloning vector pSAG1-mNeonGreen_TUB1-dTomato]
- mNeonGreen, partial [Binary vector pRATIO2131]
The peptide used to elicit this antibody is not conserved in GFP protein sequence.
This antibody is also reacting to some degree with YFP and mCherry.
Background
Product citations
Related products: mNeonGreen | Fluorescent Protein
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