Nizera ko buriwese azahora ahura nibibazo nkibi cyangwa nkibi mugihe akora reaction ya PCR, ariko ibyinshi muribyo bishobora gushyirwa mubibazo bibiri byingenzi:

Kwiyongera kwinshi kwicyitegererezo cya gene (amplification);
Byinshi cyane bitagenewe gene amplification.
Gukoresha inyongeramusaruro nimwe mungamba zisanzwe zo gukemura ibyo bibazo.Mubisanzwe uruhare rwinyongera rufite ibintu bibiri:
urwego rwa kabiriya gen (imiterere ya kabiri);
Mugabanye priming idasanzwe.
Uyu munsi, umwanditsi azakumenyesha muri make inyongeramusaruro zisanzwe mubitekerezo bya PCR n'imikorere yabo.
Inyongera zigabanya imiterere ya kabiri
sulfoxide(DMSO)
ingero za genehamwe nibirimo byinshi bya GC.Ariko, DMSO nayo igabanya cyane ibikorwa bya Taq polymerase.Kubwibyo, buriwese agomba kuringaniza inyandikorugero igerwaho nibikorwa bya polymerase.Muhinduzi atanga igitekerezo ko ushobora kugerageza kwibanda kuri DSMO itandukanye, nko kuva kuri 2% kugeza 10%, kugirango ubone kwibanda kubushakashatsi bwawe.
Imyenda idakoreshwa
Imyenda idafite ionic, nka 0.1-1% Triton X-100, Hagati ya 20 cyangwa NP-40, mubisanzwe igabanya imiterere ya kabiri ya ADN.Nubwo ibi bishobora kongera amplifisione yicyitegererezo gene, bizanatera ikibazo cyo kudasobanurwa kwihariye.Noneho, ibyo byongeweho bikora neza kubikorwa bya PCR bitanga umusaruro muke nta myanda, ariko sibyiza cyane kubitekerezo bya PRC byanduye.Iyindi nyungu yimyanda idafite ionic ni ukugabanya kwanduza SDS.Mubisanzwe mugihe cyo gukuramo ADN, SDS izazanwa ku ntambwe ya PCR, ibuza cyane ibikorwa bya polymerase.Kubwibyo, kongeraho 0.5% Hagati ya 20 cyangwa Hagati ya 40 kuri reaction irashobora gutesha agaciro ingaruka mbi za SDS.
Betaine_
Betaine irashobora kunonosora ADN igabanya imiterere yinyongera kandi mubisanzwe ni "amayobera" yiyongera kubikoresho bya PCR.Niba ushaka gukoresha betaine, ugomba gushyira betaine cyangwa betaine mono-hydrate (Betaine cyangwa Betaine mono-hydrate), ariko ntabwo hydrochloride ya betaine (Betaine HCl), ihindure kwibanda kuri 1-1.7M.Betaine irashobora kandi gufasha kunoza umwihariko kuko ikuraho ibice fatizo bigize ibice biterwa no gushonga kwa ADN / ADN.
Inyongera kugirango ugabanye priming idasanzwe
Formamide
Formamide ninyongera ikoreshwa kama PCR.Irashobora guhuza hamwe na shobuja nini hamwe nuduce duto muri ADN, bityo bikagabanya ituze rya shobuja ADN ya kabiri helix no kugabanya ubushyuhe bwa ADN.Ubwinshi bwa formamide ikoreshwa mubushakashatsi bwa PCR mubusanzwe ni 1% -5%.
Tetramethylammonium chloride( TMAC)
Tetramethylammonium chloride irashobora kongera umwihariko wa Hybridisation (hybridisation yihariye) kandi ikongerera ubushyuhe bwa ADN.Rero, TMAC irashobora gukuraho priming idasanzwe kandi ikagabanya kubeshya ADN na RNA.Niba ukoreshagutesha agaciro primersmuri reaction ya PCR, ibuka kongeramo TMAC, isanzwe ikoreshwa kuri concentration ya 15-100mM.
Ibindi Byongeweho
Usibye ibyiciro bibiri byinyongera twavuze haruguru, hari inyongeramusaruro nyinshi zisanzwe mubitekerezo bya PCR, nubwo zifite imikorere itandukanye, ningirakamaro cyane.
Magnesium ion
Magnesium ion ni cofactor yingirakamaro (cofactor) ya polymerase, nukuvuga, idafite ion ya magnesium, polymerase idakora.Nyamara, ion nyinshi za magnesium zirashobora kandi kugira ingaruka kumikorere ya polymerase.Ubwinshi bwa magnesium ion muri buri PCR reaction izatandukana.Ibikoresho bya chelating (nka EDTA cyangwa citrate), ubunini bwa DNTPs na proteyine byose bigira ingaruka kumyuka ya ion ya magnesium.Noneho, niba ufite ibibazo mubigeragezo bya PCR, urashobora kugerageza guhindura intungamubiri za magnesium zitandukanye, urugero, kuva 1.0 kugeza 4.0mM, hamwe nintera ya 0.5-1mM hagati.
Birakwiye ko tumenya ko inzinguzingo nyinshi zikonjesha zishobora kuganisha kumurongo wa chloride ya magnesium.Kubwibyo, ugomba kuyasesa burundu mbere yo kuyikoresha, ukayivanga neza mbere yo kuyikoresha.
Bovine serum albumin(Bovine albumin, BSA)
Mubushakashatsi bwa chimie ya molecular, bovine serum albumin ninyongera cyane, cyane cyane mukubuza enzyme igogora hamwe nubushakashatsi bwa PCR.Mubisubizo bya PCR, BSA ifasha mukugabanya umwanda nkibintu bya fenolike.Kandi biravugwa kandi ko bishobora kugabanya gufatira reaction kurukuta rwikizamini.Mubisubizo bya PCR, mubisanzwe kwibanda kwa BSA byongeweho bishobora kugera kuri 0.8 mg / ml.
 
Ibicuruzwa bifitanye isano:
Intwari ya PCR(hamwe n'irangi)
Intwari ya PCR