Alaabta

Cusbada Tris acetate waxaa inta badan loo isticmaalaa diyaarinta TAE (Tris-acetate-EDTA), kaas oo loo isticmaalo sida boogaha orodka iyo agarose gels.Tris Acetate-EDTA baffer waxaa loo isticmaalaa DNA agarose gel electrophoresis laakiin sidoo kale waxaa loo isticmaalaa non-denaturing RNA agarose gel electrophoresis.DNA-da laba-jibbaaran waxay u egtahay inay si degdeg ah ugu socoto TAE marka loo eego kuwa kale ee kaydiya waxaana laga yaabaa inay daalaan inta lagu jiro kororka kororka.Wareegga kaydinta ama beddelka bakhaarrada inta lagu jiro kororka elektrophoresis-ka ayaa daweyn kara awoodda wax-ka-qabashada hoose.Waxaa loo isticmaali karaa xaddi kala duwan si loo barto dhaqdhaqaaqa DNA ee xalka.Maaddaama borate ee TBE-da (Tris-Borate-EDTA Buffer, 10X Powder Pack, sc-296651) ay tahay xannibaado xooggan oo loogu talagalay enzymes badan, TAE buffer ayaa lagula talinayaa marka la eego codsiyada enzymatic ee saamiga DNA.Cusbada Tris acetate sidoo kale waa kaydka dareenka sare leh ee ATP-yada oo leh luciferase firefly.

Isticmaala: Bakhaar ordaya TAE waa kaydiyaha ugu badan ee loo isticmaalo DNA agarose gel electrophoresis, waxaana sidoo kale loo isticmaalaa RNA agarose gel electrophoresis.DNA-da laba-jibbaaran waxay u egtahay inay si degdeg ah ugu dhaqaaqdo TAE marka loo eego kuwa kale, laakiin sidoo kale waxay ku guuldareysataa inay dabbaalaan sababtoo ah yaraanta ions baffer inta lagu jiro electrophoresis dheer.Baaskiil baaskeelka ama baabuurka beddelka muddada dheer ee electrophoresis waxay magdhabi kartaa awoodda baqshiiyaha hoose.2 Kala saar kaydiyaha TAE-da uruursan si aad u hesho 1 mMTAE kayd ka kooban 40 mM Tris acetate iyo 1 mM EDTA, pH 8.3.1 mMTAE kaydinta waxaa loo isticmaali karaa labadaba agarose gels iyo sida kaydka ordaya.Xallinta ugu badan, waxaa lagu talinayaa in korantada la dabaqay uu ka hooseeyo 5V/cm (masaafada u dhaxaysa qalabka elektiroonigga ah).

Codsiga: Bakhaar ku ordaya TAE waa kaydiyaha ugu badan ee loo isticmaalo DNA agarose gel electrophoresis ee jel-ka kiimikaad, waxaana sidoo kale loo isticmaalaa kuwa aan denaturing RNA agarose gel electrophoresis.DNA-da laba-jibbaaran waxay u egtahay inay si degdeg ah ugu dhaqaaqdo TAE marka loo eego kuwa kale, laakiin sidoo kale waxay ku guuldareysataa inay dabbaalaan sababtoo ah yaraanta ions baffer inta lagu jiro electrophoresis dheer.Baaskiil baaskeelka ama baabuurka beddelka muddada dheer ee electrophoresis waxay magdhabi kartaa awoodda baqshiiyaha hoose.Xoojinta TAE-da ee xoogga leh ayaa la qasi jiray si loo helo 1 mMTAE kaydiye ka kooban 40 mM Tris acetate iyo 1 mM EDTA, pH 8.3.1 mMTAE kaydinta waxaa loo isticmaali karaa labadaba agarose gels iyo sida kaydka ordaya.Xallinta ugu badan, waxaa lagu talinayaa in korantada la dabaqay uu ka hooseeyo 5V/cm (masaafada u dhaxaysa qalabka elektiroonigga ah).

Isticmaala: Marka la ogaado ATP ee leh luciferase firefly, alaabtani waa kaydinta ugu xasaasisan;ogaanshaha xidhitaanka glutamate.

Ku-xidhka la rari karo ee [3H] l-glutamic acid ee agabka aan reseptors-ka ahayn.

[3H] L-glutamic acid oo ku xidhan tuubooyinka microfuge iyo dhalooyinka ayaa lagu baadhay afar kayd.Ku-xidhka asalka ah ee walxahan ayaa ahaa mid dayacan, laakiin waxaa kordhay centrifugation ama nuugista Tris-HCl iyo Tris-citrate buffer.Xiritaankani aad buu u yaraa ama Markii HEPES-KOH, ama Tris-acetate buffer la isticmaalay beddelkeeda.[3H] L-glutamate ku xidhida tuubooyinka microfuge waxaa hakiyay L-laakin ma aha D-isomers of glutamate iyo aspartate.DL-2-amino-7-phosphonoheptanoic acid sidoo kale ma joojin xiritaanka.Xeryahooda kale ee muujiyay xannibaad hoose ama dhexdhexaad ah waxay ahaayeen: N-methyl-D-aspartate, qusqualate, L-glutamic acid diethyl ester, N-methyl-L-aspartate, kainate, iyo 2-amino-4-phosphonobutyrate.Isku xidhka waxaa hakiyay xuubabka maskaxda ee jiirka oo go'ay.Isku-xidhka borotiinka-ku-tiirsanaanta [3H] glutamate ayaa lagu helay diyaarinta xuub barafaysan oo soo noqnoqda markii xidhidhisku lagu sameeyay bakhaar Tris-acetate.Waxaa lagu talinayaa in Tris-acetate ama HEPES-KOH kaydinta loo isticmaalo glutamate bin ding assay.Haddii Tris-HCl ama Tris-citrate baffer la isticmaalo, tijaabinta xakamaynta ku haboon waa in la sameeyaa si loo saxo xidhitaanka tuubooyinka microfuge ama filtarrada galaaska.