関: 5-bromo-4-chloro-3-indolyl-beta-D-galactoside

WordNet

a unit of gravitational acceleration equal to one centimeter per second per second (named after Galileo)
alliterative term for girl (or woman)
the 24th letter of the Roman alphabet (同)x, ex
a unit of force equal to the force exerted by gravity; used to indicate the force to which a body is subjected when it is accelerated (同)gee, g-force

PrepTutorEJDIC

女の子,少女(girl)
Christ / Christian
x-axis

Wikipedia preview

出典(authority):フリー百科事典『ウィキペディア（Wikipedia）』「2015/09/08 12:22:55」(JST)

wiki ja

5-ブロモ-4-クロロ-3-インドリル-β-D-ガラクトピラノシド、通称X-gal（エックスギャル）は、ガラクトースと置換インドールから構成される有機化合物である。分子生物学において、β-ガラクトシダーゼにより分解されると青い色を呈する性質を利用して、組換えプラスミドに遺伝子が挿入されたことを確認する簡便な検査法である、ブルーホワイトスクリーニングに用いられている。

作用機構

酵母あるいは大腸菌において、ラクトースオペロンを構成する遺伝子lacZの翻訳産物であるβ-ガラクトシダーゼが発現すると、X-galはガラクトースと5-ブロモ-4-クロロ-3-インドールに切断される。5-ブロモ-4-クロロ-3-インドールは酸化されて、不溶性の青い色素である5,5'-ジブロモ-4,4'-ジクロロ-インディゴに変化する。これを用いて、寒天培地の中にX-galとβ-ガラクトシダーゼの誘導剤（通常はイソプロピル-β-チオガラクトピラノシド（IPTG））を加えておくことにより、ラクトースオペロンを発現しているコロニーを容易に見分けることができる。

応用

レポーターアッセイ

lacZ遺伝子はX-galを含む培地組み合わせてレポーター遺伝子として用いられる。例えば、ツーハイブリッド法では、酵母やバクテリア内で、タンパク質間相互作用が成立したものとしていないものを区別する必要がある。その際、プロモーターにの下流にlacZ遺伝子を置くと、コロニーが青色を呈したことでタンパク質間相互作用が成立したことを確認することが出来る^[1]。この方法の適用は、コロニーの数が約10⁶より少なく、コロニー同士を区別できる時に限られる^[1]。X-galが分解されると、インドールの不快な匂いがする。

飲料水の検査

分子生物学における使用に加え、X-GalはE. coliや大腸菌群が飲料水に含まれているかどうかを調べるのに用いられる^[2]。

脚注

^ ^a ^b Joung, J.; Ramm, E.; Pabo, C. (2000). "A bacterial two-hybrid selection system for studying protein-DNA and protein-protein interactions". Proc. Natl. Acad. Sci., USA 97 (13): 7382–7387. doi:10.1073/pnas.110149297. PMC 16554. PMID 10852947.
^ 古畑勝則 (2002). "飲料水の大腸菌群検査法" (PDF). イーズ 27.

この項目は、生物学に関連した書きかけの項目です。この項目を加筆・訂正などしてくださる協力者を求めています（プロジェクト:生命科学／Portal:生物学）。

wiki en

[Wiki en表示]

X-gal (also abbreviated BCIG for 5-bromo-4-chloro-3-indolyl-β-D-galactopyranoside) is an organic compound consisting of galactose linked to a substituted indole. The compound was synthesized by Jerome Horwitz and collaborators in Detroit, MI, in 1964.^[1] The formal chemical name is often shortened to less accurate but also less cumbersome phrases such as bromochloroindoxyl galactoside. The X from indoxyl may be the source of the X in the X-gal contraction. X-gal is much used in molecular biology to test for the presence of an enzyme, β-galactosidase. It is also used to detect activity of this enzyme in histochemistry and bacteriology. X-gal is one of many indoxyl glycosides and esters that yield insoluble blue compounds similar to indigo as a result of enzyme-catalyzed hydrolysis.^[2]

Uses

X-gal is an analog of lactose, and therefore may be hydrolyzed by the β-galactosidase enzyme which cleaves the β-glycosidic bond in D-lactose. X-gal, when cleaved by β-galactosidase, yields galactose and 5-bromo-4-chloro-3-hydroxyindole. The latter then spontaneously dimerizes and is oxidized into 5,5'-dibromo-4,4'-dichloro-indigo, an intensely blue product which is insoluble. X-gal itself is colorless, the presence of blue-colored product therefore may be used as a test for the presence of an active β-galactosidase. This easy identification of an active enzyme allows the gene for β-galactosidase (the lacZ gene) to be used as a reporter gene in various applications.^[3]

Reaction

Cloning

In gene cloning, X-gal is used as a visual indication of whether a cell expresses a functional β-galactosidase enzyme in a technique called blue/white screening. This method of screening is a convenient way of distinguishing a successful cloning product from other unsuccessful ones.

The blue/white screening method relies on the principle of α-complementation of the β-galactosidase gene, where a fragment of the lacZ gene (lacZα) in the plasmid can complement another mutant lacZ gene (lacZΔM15) in the cell. Both genes by themselves produce non-functional peptides, however, when expressed together, as when a plasmid containing lacZα is transformed into a lacZΔM15 cells, they form a functional β-galactosidase. The presence of an active β-galactosidase may be detected when cells are grown in plates containing X-gal, the blue-colored product precipitated within cells resulted in the characteristic blue colonies. However, the multiple cloning site, where a gene of interest may be ligated into the plasmid vector, is located within the lacZα gene. Successful ligation therefore disrupts the lacZα gene, α-complementation is therefore also disrupted and no functional β-galactosidase can form, resulting in white colonies. Cells containing successfully ligated insert can then be easily identified by its white coloration from the unsuccessful blue ones. Example of cloning vectors used for this test are pUC19, pBluescript, pGem-T Vectors, and it also requires the use of specific E. coli host strains such as DH5α which carries the mutant lacZΔM15 gene.

Variants

X-gal has a number of variants, which are similar molecules with slight differences serving mainly to produce colors other than blue as a signal.

Short name	Long name	Color
Blue-Gal, Bluo-Gal	5-Bromo-3-indolyl β-D-galactopyranoside	Dark blue^[4]
Rose-Gal, Salmon-Gal, Y-Gal, Red-Gal	6-Chloro-3-indolyl-β-D-galactopyranoside	Pink^[5]
Purple-β-D-Gal	5-Iodo-3-indolyl-β-D-galactopyranoside	Purple^[6]
Magenta glucoside, Magenta-GLC, Magenta gal	5-Bromo-6-chloro-3-indolyl-β-D-glucopyranoside	Magenta^[7]
Green-β-D-gal	N-Methylindolyl-β-D-galactopyranoside	Green^[8]
MUG, MUGA	4-Methylumbelliferyl β-D-galactopyranoside	Fluorescent^[9] (λ_ex = 365, λ_em = 455)

Protein-protein interactions

In two-hybrid analysis, β-galactosidase may be used as a reporter to identify proteins that interact with each other. In this method, genome libraries may be screened for protein interaction using yeast or bacterial system. Where there is a successful interaction between proteins being screened, it will result to the binding of an activation domain to a promoter. If the promoter is linked to a lacZ gene, the production of β-galactosidase, which results in the formation of blue-pigmented colonies in the presence of X-gal, will therefore indicate a successful interaction between proteins.^[10] This technique may be limited to screening libraries of size of less than around 10⁶.^[10] The successful cleavage of X-gal also creates a noticeably foul odor due to the volatilization of indole.

References

^ Horwitz JP and 7 others, 1964. Substrates for cytochemical demonstration of enzyme activity. I. Some substituted 3-indolyl-β-D-glycopyranosides. Journal of Medicinal Chemistry 7: 574-575.
^ Kiernan JA 2007. Indigogenic substrates for detection and localization of enzymes. Biotechnic & Histochemistry 82(2): 73-103.
^ Sandhu, Sardul Singh (2010). Recombinant DNA Technology. I K International Publishing House. p. 116. ISBN 978-9380578446.
^ "5-Bromo-3-indolyl β-D-galactopyranoside". Retrieved 4 February 2014.
^ "Salmon-Gal - PubChem". Retrieved 4 February 2014.
^ "Purple-beta-D-Gal - PubChem". Retrieved 4 February 2014.
^ "5-Bromo-6-chloro-3-indolyl-β-D-glucopyranoside". Retrieved 4 February 2014.
^ "Green-β-D-Gal - Biotium, Inc.". Retrieved 4 February 2014.
^ "4-Methylumbelliferyl β-D-galactopyranoside". Retrieved 4 February 2014.
^ ^a ^b Joung J, Ramm E, Pabo C (2000). "A bacterial two-hybrid selection system for studying protein-DNA and protein-protein interactions". Proc Natl Acad Sci USA 97 (13): 7382–7. doi:10.1073/pnas.110149297. PMC 16554. PMID 10852947.

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★リンクテーブル★

リンク元	「5-bromo-4-chloro-3-indolyl-beta-D-galactoside」「5-ブロモ-4-クロロ-3-インドリル-β-D-ガラクトシド」
関連記事	「gal」「X」「g」

「5-bromo-4-chloro-3-indolyl-beta-D-galactoside」

X-gal
Names
	IUPAC name 5-bromo-4-chloro-3-indolyl-β-D-galactopyranoside
Identifiers
CAS Registry Number	7240-90-6
ChEBI	CHEBI:75055
ChemSpider	58680
	InChI InChI=1S/C14H15BrClNO6/c15-5-1-2-6-9(10(5)16)7(3-17-6)22-14-13(21)12(20)11(19)8(4-18)23-14/h1-3,8,11-14,17-21H,4H2/t8-,11+,12+,13-,14-/m1/s1 ; Key: OPIFSICVWOWJMJ-AEOCFKNESA-N ; InChI=1/C14H15BrClNO6/c15-5-1-2-6-9(10(5)16)7(3-17-6)22-14-13(21)12(20)11(19)8(4-18)23-14/h1-3,8,11-14,17-21H,4H2/t8-,11+,12+,13-,14-/m1/s1; Key: OPIFSICVWOWJMJ-AEOCFKNEBZ ;
Jmol-3D images	Image
MeSH	X-gal
PubChem	65181
	SMILES Brc3ccc2c(c(O[C@@H]1O[C@@H]([C@H](O)[C@H](O)[C@H]1O)CO)cn2)c3Cl ;
Properties
Chemical formula	C14H15BrClNO6
Molar mass	408.629
	Except where otherwise noted, data are given for materials in their standard state (at 25 °C [77 °F], 100 kPa).
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	Infobox references

X-gal
	IUPAC名 5-bromo-4-chloro-3-indolyl- beta-D-galactopyranoside
識別情報
CAS登録番号	7240-90-6
PubChem	65181
日化辞番号	J212.257J
MeSH	X-gal
	SMILES C1=CC(=C(C2=C1NC=C2O[C@H]3[C@@H]([C@H]([C@H]([C@H](O3)CO)O)O)O)Cl)Br;
特性
化学式	C14H15BrClNO6
モル質量	408.63 g mol−1
外観	固体
危険性
MSDS	Science Lab
	特記なき場合、データは常温 (25 °C)・常圧 (100 kPa) におけるものである。