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GREEN STAIN DNA/RNA DETECTION

Introduction

GREEN STAIN (10,000X in DMSO) is a versatile and easy-to-use tool for the detection of nucleic acids (DNA and RNA) in electrophoretic gels.

Features

  • Three easy to use protocols: pre-electrophoresis gel staining for the highest sensitivity; sample pre-staining for the best migration results; post-electrophoresis gel staining for particular protocol needs.
  • Very low toxicity.

INTRODUCTION

Nucleic acid gel staining has been performed with ethidium bromide for decades.
Even though ethidium bromide is very economic and easy to use, its mutagenicity and hazardous waste disposal fueled the development of DNA gel staining alternatives. Cyanagen has developed GREEN STAIN to solve these issues.
GREEN STAIN has a high binding affinity for nucleic acids providing bright green emitting DNA-dye complexes. It preferentially binds double-strand DNA and also allows single-strand DNA and RNA detection, although with lower sensitivity. Bright green fluorescent bands and very low background fluorescence are its major features. The nucleic acid-bound GREEN STAIN is efficiently excited at ~254 nm and ~488 nm; it can also be excited at ~312 nm with loss of sensitivity. Detection can be performed with the same instruments used for ethidium bromide and SYBR® Green gel staining such as standard UV transilluminator (254 nm) as well as with CCD-camera imaging system or laser-based scanner selecting the SYBR® Green filter. GREEN STAIN can be used for any gel staining application requiring the detection of up to 250 pg of genomic dsDNA with sufficient sensitivity with 312 and 254 nm transillumination (up to ~ 100 pg with 460 nm epillumination).

GREEN STAIN can be used with one of the following protocols:
- pre-electrophoresis gel staining for the highest sensitivity (up to 100 pg)
- sample pre-staining for the best migration results with a loss of sensitivity (500 pg)
- post-electrophoresis gel staining for particular protocol needs

 

Excitation/emission spectra of Green Stain bound to dsDNA Excitation/emission spectra of GREEN STAIN bound to dsDNA

 

COMPARISON WITH OTHER METHODS

 
Detection Limit: pre-electrophoresis gel staining

Detection Limit: pre-electrophoresis gel staining

The nucleic acid-bound GREEN STAIN is efficiently excited at ~254 nm and ~488 nm. It can also be excited at 312 nm with loss of sensitivity (at least 5-6 times). When using UV transilluminators at 312 nm we recommend to increase the exposure time.

Genomic DNA was electrophoresed on a 1% agarose gel stained with a 1:10000 diluition of GREEN STAIN (pre-electrophoresis gel staining). Gel visualization was done with (A) a standard UV-transilluminator (254 nm), exposure time 1,6 sec; (B) GE Healthcare Life Sciences "ImageQuant LAS 4000“ (312 nm), exposure time 5 sec; (C) GE Healthcare Life Sciences "ImageQuant LAS 4000" (460 nm), exposure time 2 sec.

 
Detection Limit: sample pre-staining protocol

Detection Limit: sample pre-staining protocol

The nucleic acid-bound GREEN STAIN is efficiently excited at ~254 nm and ~488 nm. It can also be excited at 312 nm with loss of sensitivity (at least 5-6 times). When using UV transilluminators at 312 nm we recommend to increase the exposure time.

Genomic DNA was electrophoresed on a 1% agarose. Samples were stained with GREEN STAIN 500X (sample pre-staining protocol). Gel visualization was done with (A) a standard UV-transilluminator (254 nm), exposure time 1,6 sec; (B) GE Healthcare Life Sciences "ImageQuant LAS 4000“ (312 nm), exposure time 5 sec; (C) GE Healthcare Life Sciences "ImageQuant LAS 4000" (460 nm), exposure time 2 sec.

 
DNA Ladders: pre-electrophoresis gel staining

DNA Ladders: pre-electrophoresis gel staining

A) 30 ng of GeneRuler™ 1 Kb Plus DNA Ladder (Thermo Scientifc) was electrophoresed on a 1% agarose gel at 100 V for 60 minutes.

B) 30 ng of GeneRuler™ 100 bp Plus DNA Ladder (Thermo Scientifc) was electrophoresed on a 2% agarose gel at 100 V for 60 minutes.

C) 30 ng of GeneRuler™ Low range DNA Ladder (Thermo Scientifc) was electrophoresed on a 3% agarose gel at 100 V for 60 minutes.

All the gels were stained with a 1:10000 diluition of GREEN STAIN (pre-electrophoresis gel staining).
Gel visualization was done with GE Healthcare Life Sciences "ImageQuant LAS 4000" (460 nm flter), exposure time 2 sec.

   
DNA Ladders: sample pre-staining

DNA Ladders: sample pre-staining

A) 500 ng of GeneRuler™ 1 Kb Plus DNA Ladder (Thermo Scientifc) was electrophoresed on a 1% agarose gel at 100 V for 60 minutes.

B) 500 ng of GeneRuler™ 100 bp Plus DNA Ladder (Thermo Scientifc) was electrophoresed on a 2% agarose gel at 100 V for 60 minutes.

C) 500 ng of GeneRuler™ Low range DNA Ladder (Thermo Scientifc) was electrophoresed on a 3% agarose gel at 100 V for 60 minutes.

All the samples were stained with GREEN STAIN 1:200 (sample -staining).
Gel visualization was done with GE Healthcare Life Sciences "ImageQuant LAS 4000" (460 nm flter), exposure time 2 sec.

   
254 nm transillumination: comparison with competitors

254 nm transillumination: comparison with competitors

Genomic DNA was electrophoresed on a 1% agarose gel stained with a 1:10000 diluition of the dye indicated on the right (preelectrophoresis gel staining). Gel visualization was done with a standard UV-transilluminator (254 nm), exposure time 1,6 sec.

   
312 nm transillumination: comparison with competitors

312 nm transillumination: comparison with competitors

Genomic DNA was electrophoresed on a 1% agarose gel stained with a 1:10000 diluition of the dye indicated on the right (preelectrophoresis gel staining). Gel visualization was done with GE Healthcare Life Sciences "ImageQuant LAS 4000“ (312 nm), exposure time 5 sec.

   
460 nm epillumination: comparison with competitors

460 nm epillumination: comparison with competitors

Genomic DNA was electrophoresed on a 1% agarose gel stained with a 1:10000 diluition of the dye indicated on the right (preelectrophoresis gel staining). Gel visualization was done with GE Healthcare Life Sciences "ImageQuant LAS 4000" (460 nm), exposure time 2 sec.

 
   
Comparison with Ethidium Bromide

Comparison with Ethidium Bromide

Genomic DNA was electrophoresed on a 1% agarose gel stained with a 1:10000 diluition of the dye indicated on the left (preelectrophoresis gel staining). Gel visualization was done with a standard UV-transilluminator (254 nm), exposure time 1,6 sec; GE Healthcare Life Sciences "ImageQuant LAS 4000“ (312 nm), exposure time 5 sec.

   
Comparison with SYBR® Safe

Comparison with SYBR® Safe

Genomic DNA was electrophoresed on a 1% agarose gel stained with a 1:10000 diluition of the dye indicated on the left (preelectrophoresis gel staining). Gel visualization was done with a standard UV-transilluminator (254 nm), exposure time 1,6 sec; GE Healthcare Life Sciences "ImageQuant LAS 4000“ (312 nm), exposure time 5 sec; GE Healthcare Life Sciences "ImageQuant LAS 4000" (460 nm), exposure time 2 sec.

   
Comparison with SYBR® Green I

Comparison with SYBR® Green I

Genomic DNA was electrophoresed on a 1% agarose gel stained with a 1:10000 diluition of the dye indicated on the left (preelectrophoresis gel staining). Gel visualization was done with a standard UV-transilluminator (254 nm), exposure time 1,6 sec; GE Healthcare Life Sciences "ImageQuant LAS 4000“ (312 nm), exposure time 5 sec; GE Healthcare Life Sciences "ImageQuant LAS 4000" (460 nm), exposure time 2 sec.

   
Comparison GelRed™

Comparison with GelRed™

Genomic DNA was electrophoresed on a 1% agarose gel stained with a 1:10000 diluition of the dye indicated on the left (preelectrophoresis gel staining). Gel visualization was done with a standard UV-transilluminator (254 nm), exposure time 1,6 sec; GE Healthcare Life Sciences "ImageQuant LAS 4000“ (312 nm), exposure time 5 sec.

   
Comparison with GelGreen™

Comparison with GelGreen™

Genomic DNA was electrophoresed on a 1% agarose gel stained with a 1:10000 diluition of the dye indicated on the left (preelectrophoresis gel staining). Gel visualization was done with a standard UV-transilluminator (254 nm), exposure time 1,6 sec; GE Healthcare Life Sciences "ImageQuant LAS 4000“ (312 nm), exposure time 5 sec; GE Healthcare Life Sciences "ImageQuant LAS 4000" (460 nm), exposure time 2 sec.