DEPC-treated (and therefore RNase-free) water is used in handling of RNA in the laboratory to reduce the risk of RNA being degraded by RNases.
Water is usually treated with 0.1% v/v DEPC for at least 2 hours at 37 °C and then autoclaved (at least 15 min) to inactivate traces of DEPC. Inactivation of DEPC in this manner yields CO2 and ethanol. Higher concentrations of DEPC are capable of deactivating larger amounts of RNase, but remaining traces or byproducts may inhibit further biochemical reactions such as in vitro transcription. Furthermore, chemical modification of RNA such as carboxymethylation is possible when traces of DEPC or its byproducts are present, resulting in impaired recovery of intact RNA even after buffer exchange (after precipitation).
DEPC treated water for use in a laboratory
DEPC is unstable in water and susceptible to hydrolysis to carbon dioxide and ethanol, especially in the presence of a nucleophile. For this reason, DEPC cannot be used with Tris or HEPES buffers. In contrast, it can be used with phosphate-buffered saline or MOPS.[3] A handy rule is that enzymes or chemicals which have active -O:, -N: or -S: cannot be treated with DEPC to become RNase-free, as DEPC reacts with these species. Furthermore, DEPC degradation products can inhibit in vitro transcription.
DEPC derivatization of histidines is also used to study the importance of histidyl residues in enzymes. Modification of histidine by DEPC results in carbethoxylated derivates at the N-omega-2 nitrogen of the imidazole ring. DEPC modification of histidines can be reversed by treatment with 0.5 M hydroxylamine at neutral pH.
DEPC can also be used for probing the structure of double-stranded DNA.[citation needed]
^Chirgwin, John M; et al. (1979). "Isolation of biologically active ribonucleic acid from sourcesenriched in ribonuclease". Biochemistry. 18 (24): 5294–5299. doi:10.1021/bi00591a005. PMID 518835.
^Wolf, Barry; Lesnaw, Judith A.; Reichmann, Manfred E. (1970). "A Mechanism of the Irreversible Inactivation of Bovine Pancreatic Ribonuclease by Diethylpyrocarbonate. A General Reaction of Diethylpyrocarbonate with Proteins". European Journal of Biochemistry. 13 (3): 519–25. doi:10.1111/j.1432-1033.1970.tb00955.x. PMID 5444158.
^"FAQ about DEPC". Sigma-Aldrich. Retrieved 12 August 2012.
External linksedit
DEPC at OpenWetWare life scientist wiki
RNase and DEPC Treatment
April 10, 2024
diethyl, pyrocarbonate, depc, also, called, diethyl, dicarbonate, iupac, name, used, laboratory, inactivate, rnase, enzymes, water, laboratory, utensils, does, covalent, modification, histidine, most, strongly, lysine, cysteine, tyrosine, residues, namesprefer. Diethyl pyrocarbonate DEPC also called diethyl dicarbonate IUPAC name is used in the laboratory to inactivate RNase enzymes in water and on laboratory utensils It does so by the covalent modification of histidine most strongly lysine cysteine and tyrosine residues 1 2 Diethyl pyrocarbonate NamesPreferred IUPAC name Diethyl dicarbonateOther names DiethylpyrocarbonateDiethyl oxydiformateEthoxyformic anhydridePyrocarbonic acid diethyl esterDEPCIdentifiersCAS Number 1609 47 8 Y3D model JSmol Interactive imageChEBI CHEBI 59051 YChEMBL ChEMBL55517 YChemSpider 2943 YECHA InfoCard 100 015 039EC Number 216 542 8KEGG C11592 YMeSH DiethylpyrocarbonatePubChem CID 3051UNII LMR3LZG146 YCompTox Dashboard EPA DTXSID2051764InChI InChI 1S C6H10O5 c1 3 9 5 7 11 6 8 10 4 2 h3 4H2 1 2H3 YKey FFYPMLJYZAEMQB UHFFFAOYSA N YInChI 1 C6H10O5 c1 3 9 5 7 11 6 8 10 4 2 h3 4H2 1 2H3Key FFYPMLJYZAEMQB UHFFFAOYAUSMILES O C OCC OC O OCCPropertiesChemical formula C 6H 10O 5Molar mass 162 141 g mol 1Appearance Clear colorless liquidDensity 1 101 g mL at 25 C1 121 g mL at 20 CBoiling point 93 to 94 C 199 to 201 F 366 to 367 K at 24 hPaHazardsOccupational safety and health OHS OSH Main hazards HarmfulGHS labelling PictogramsSignal word WarningHazard statements H302 H315 H319 H332 H335Precautionary statements P261 P264 P270 P271 P280 P301 P312 P302 P352 P304 P312 P304 P340 P305 P351 P338 P312 P321 P330 P332 P313 P337 P313 P362 P403 P233 P405 P501Flash point 69 C 156 F 342 K closed cupLethal dose or concentration LD LC LD50 median dose Oral rat 850 mg kgRelated compoundsRelated compounds Di tert butyl dicarbonateDimethyl dicarbonateExcept where otherwise noted data are given for materials in their standard state at 25 C 77 F 100 kPa Y verify what is Y N Infobox references DEPC treated and therefore RNase free water is used in handling of RNA in the laboratory to reduce the risk of RNA being degraded by RNases Water is usually treated with 0 1 v v DEPC for at least 2 hours at 37 C and then autoclaved at least 15 min to inactivate traces of DEPC Inactivation of DEPC in this manner yields CO2 and ethanol Higher concentrations of DEPC are capable of deactivating larger amounts of RNase but remaining traces or byproducts may inhibit further biochemical reactions such as in vitro transcription Furthermore chemical modification of RNA such as carboxymethylation is possible when traces of DEPC or its byproducts are present resulting in impaired recovery of intact RNA even after buffer exchange after precipitation DEPC treated water for use in a laboratoryDEPC is unstable in water and susceptible to hydrolysis to carbon dioxide and ethanol especially in the presence of a nucleophile For this reason DEPC cannot be used with Tris or HEPES buffers In contrast it can be used with phosphate buffered saline or MOPS 3 A handy rule is that enzymes or chemicals which have active O N or S cannot be treated with DEPC to become RNase free as DEPC reacts with these species Furthermore DEPC degradation products can inhibit in vitro transcription DEPC derivatization of histidines is also used to study the importance of histidyl residues in enzymes Modification of histidine by DEPC results in carbethoxylated derivates at the N omega 2 nitrogen of the imidazole ring DEPC modification of histidines can be reversed by treatment with 0 5 M hydroxylamine at neutral pH DEPC can also be used for probing the structure of double stranded DNA citation needed See also editDECReferences edit Chirgwin John M et al 1979 Isolation of biologically active ribonucleic acid from sourcesenriched in ribonuclease Biochemistry 18 24 5294 5299 doi 10 1021 bi00591a005 PMID 518835 Wolf Barry Lesnaw Judith A Reichmann Manfred E 1970 A Mechanism of the Irreversible Inactivation of Bovine Pancreatic Ribonuclease by Diethylpyrocarbonate A General Reaction of Diethylpyrocarbonate with Proteins European Journal of Biochemistry 13 3 519 25 doi 10 1111 j 1432 1033 1970 tb00955 x PMID 5444158 FAQ about DEPC Sigma Aldrich Retrieved 12 August 2012 External links editDEPC at OpenWetWare life scientist wiki RNase and DEPC Treatment Retrieved from https en wikipedia org w index php title Diethyl pyrocarbonate amp oldid 1172273902, wikipedia, wiki, book, books, library,
