- X -Gal
Colorless crystalline solid
225-230 ° C
Partially soluble in water
Template: Infobox chemical / molecular formula search available
5-bromo -4-chloro -3- indoxyl - β -D-galactopyranoside, and X -Gal, is an artificial glycoside and a chromogenic substrate for the enzyme β -galactosidase.
The enzyme β -galactosidase X-gal is hydrolyzed to galactose and 5-bromo- 4-chloro- indoxyl (5- bromo-4- chloro-3- hydroxyindole ). The 5 -bromo -4-chloro - indoxyl is, 4' -dichloro- indigo oxidized by oxygen in the air to deep blue dye 5,5 '- dibromo -4.
In biochemistry, X-Gal is used for the qualitative determination of the activity of β -galactosidase. For the quantitative determination, however, the substrate o -nitrophenyl- β -D-galactopyranoside (ONPG ) is used.
In molecular biology, X -Gal is used for blue - white selection. In many cloning vectors lies in the amino-terminal coding sequence of the β -galactosidase ( α - fragment), a multiple cloning site can be introduced into the foreign DNA, such as PCR fragments. Thus, the gene sequence of the β -galactosidase is interrupted and the enzyme is not expressed. Consequently, the X-gal is not cleaved and thus not a blue dye. The resulting colonies after transformation then appear not as blue as the empty vector, but white.
X-Gal is often (including lac operon induction of the lac promoter, see ) are used for the screening of blue colonies in conjunction with IPTG. It is also used for the screening of β -galactosidase reporter gene activity in transfections of eukaryotic cells and for the detection of β -galactosidase in immunology and histochemical applications.
Other chromogenic substrates:
- 5-bromo -3-indolyl - β -D-galactopyranoside ( Blue -Gal, Bluo -gal)
- 6-chloro -3-indolyl - β -D -galactopyranoside (Y- Gal, Red- Gal, Gal - Rose, Salmon -gal)
- 5-iodo -3-indolyl - β -D-galactopyranoside ( Gal - purple )
- 5-bromo -6-chloro -3-indolyl - β -D-galactopyranoside ( Gal - magenta )
- N- methylindolyl - β -D-galactopyranoside ( Gal - Green )
Other fluorogenic substrates:
- 4-methyl- umbelliferyl - β -D-galactopyranoside (MUG ) ( λex = 365 nm, λem = 455 nm)