Comparison of transfection efficiencies of a few in vitro DNA transfection reagents: GeneExpresso, LipoExpress, Lipofectamine 2000, and Expressfect

GeneExpresso and LipoExpress are from Lab Supply Mall
Lipofectamine 2000 is from Invitrogen.

Expressfect is from Denville Scientific, Inc. (http://www.denvillescientific.com), Expressfect is an OEM product (SoFast) from China (http://www.sunmabio.com, http://www.actgene.com/Convoy.htm).

Lipofectamine 2000	LipoExpress
GeneExpresso	Expressfect (SoFast)
Efficiency comparison of GeneExpresso, LipoExpress, Lipofectamine 2000, and Expressfect on Hela cells. A nuclear targeted GFP DNA was transfected with GeneExpresso, LipoExpress, Lipofectamine 2000, and Expressfect respectively per manufacturer’s protocols. The efficiency was checked 48 hours post transfection.
FACS Analysis

Conclusion:
Both GeneExpresso and LipoExpress have higher transfection efficiency than Lipofectamine 2000. Expressfect (OEM of SoFast) should not be used as transfection reagent.

GRGreen is a nucleic acid stain for detecting nucleic acids, e.g., double-stranded DNA, in agarose gel. It can be used for replacing mutagenic ethidium bromide (EB).   

Compared to EB which is a very strong mutagen, GRGreen caused fewer mutations than EB in the Ames test.

• Available at 10,000X in H₂O for better safety.
• Compatible with UV or blue light transilluminator and common gel documentation systems.
• Will not affect downstream experiments: compatible with all gel purification kits tested, will not inhibit ligation reaction etc.
• Compatible with Sodium Borate Electrophoresis Buffer: Run gel 2-3 times faster at higher voltage, resolve shaper bands in minutes, and less heat generation.
• Cut out DNA bands for subclonning under safer blue light: No mutations caused by EB and UV light.

Stain A	GRGreen
DarkReader Blue Light Transilluminator
Stain A	GRGreen
UV Transilluminator

 Fig. 2, GRGreen in water is more sensitive than Stain A in water.

1% agarose gel prepared with SeaKem LE Agarose (Lonza) and 1X GRGreen or 1X Stain A (from competitor A), run with 1X Lithium Borate Fast Electrophoresis Buffer at 100 V.

Left: 100 bp DNA ladder, 250ng, 125 ng, 62.5 ng and 31.3 ng, total ng DNA per lane.

Right: 1kb DNA ladder, 250ng, 125 ng, 62.5 ng and 31.3 ng, total ng DNA per lane.

TAE	TBE	Sodium Borate

Fig. 3, GRGreen is compatible with three common running buffers.

1% agarose gel prepared with SeaKem LE Agarose (Lonza) and 1X GRGreen, run with 1X  three Electrophoresis Buffers at 100 V.

Storage: Store at 4 ^oC.

Protocol:

1. Prepare 20 to 40 ml of agarose gel solution (concentration from 0.7~2.0%) with TAE, TBE or Borate Buffer in a 250 ml flask and mix it thoroughly. Place the flask in the microware, heat on high until the solution is completely clear and no small floating particles are visible (about 2~3 minutes).

2. After the gel solution cool to about 55 oC, add GRGreen to the solution to 1X final concentration. Swirl the flask gently to mix the solution and avoid forming bubbles.

3. Pour the gel solution into a gel tray until the comb teeth are immersed about 1/4~1/2 into the gel solution.

4. After the agarose gel has solidified you can perform electrophoresis using either 0.5 to 1X TAE, TBE or Borate Buffer.

5. Detect the bands using UV or blue light transilluminator.

FAQ

1, Should I wear gloves when using this dye?

   You should exercise common safe laboratory practices when using this reagent.

2, What blue light transilluminator should I used with GRGreen dye?

    DarkReader.

3, What filter should I use for blue light transilluminator?

    Filter from Clare Chemical.

 MSDS: GRGreen MSDS, Disposal Guide

Protocol

This Perl script shows you how to automate data download from a website.

#!/usr/bin/perl
use strict;
# load LWP library:
#use LWP::Simple;
#use LWP::RobotUA;
use HTML::Parse;
use WWW::Mechanize;

#my $agent=’Mozilla/4.0 (compatible; MSIE 8.0; Windows NT 5.1; .NET CLR 1.0.3705; .NET CLR 2.0.50727)’;

my $mech = WWW::Mechanize->new();
$mech->agent_alias( ‘Windows IE 6′ );

my $url = ‘http://www.xxx.com/catalog/index.php?_a=viewProd&productId=’;

my $i=1600;

for (my $count = $i; $count >= 1; $count–)
{
    my $new_url=$url. $count;
    #my $content = get $new_url;
    $mech->get( $new_url );
    my $output_page = $mech->content();

    #my $parsed_html = HTML::Parse::parse_html($content);

    if($output_page !~ m/That product could not be found/i)
    {
        print “downloading $count\n”;
       
        open (OUT, “>$count.html”) or die “—\n”;  
        print OUT $output_page;
      close (OUT);
  }

}

1, Open a shell then type

sudo perl -MCPAN -we ‘shell’

2, install the module

cpan> install WWW::Mechanize
…
…
/usr/bin/make install — OK

cpan> q

1, Stop MySQL server

 /etc/init.d/mysqld stop

2, Backup your databases.

cp /database_dir/database_files /backup_dir/.

3,  Remove Old MySQL files

yum remove mysql

4, Reinstall database

yum install mysql-server mysql

5, start database

/etc/init.d/mysqld start

6, create new users

mysql –user=root mysql -p
Enter password:

mysql> GRANT ALL PRIVILEGES ON *.* TO ‘admin_user’@'%my_IP_address’ IDENTIFIED BY ‘only_FBI_knows’ WITH GRANT OPTION;

mysql> quit;

/etc/init.d/mysqld stop

7, edit my.cnf file

    datadir=/disk2/mysql_data

copy files to data directory

 cp -R /var/lib/mysql/* /disk2/mysql_data/.
 cp -R /backup_dir/* /disk2/mysql_data/.
 chown -R mysql:mysql  /disk2/mysql_data

8, restart mysql

/etc/init.d/mysqld start
or
/sbin/service mysqld start

9, Import backup data

mysql -p -h localhost labsupply < labsupply.sql

data  backup command:
/usr/bin/mysqldump -u root –password=topsecret –databases labsupply > labsupply.sql

10, Make sure MySQL will be started on reboot:

chkconfig mysqld on