Universal SYBR GREEN qPCR Premix (Blue)

Nambala ya mphaka: HCB5041B

Universal Blue qPCR Master Mix (Dye Based) ndi yankho lakale la 2 × real-time quantitative quantitative PCR yodziwika ndi kukhudzika kwakukulu komanso kusiyanasiyana, ndi yabuluu mumtundu, ndipo imakhala ndi zotsatira za kutsata zitsanzo.Chigawo chapakati cha Taq DNA polymerase chimagwiritsa ntchito ma antibody otentha poyambira kuti alepheretse kukulitsa kosagwirizana kwenikweni chifukwa cha kuyambika koyambira pokonzekera zitsanzo.Nthawi yomweyo, chilinganizocho chimawonjezera zinthu zomwe zimalimbikitsa kukulitsa mphamvu ya PCR ndikufananiza kukulitsa kwa majini okhala ndi GC zosiyanasiyana (30 ~ 70%), kuti PCR yochulukira ikhoza kupeza ubale wabwino wamzere mu kuchuluka kwakukulu. dera.Izi zili ndi ROX Passive Reference Dye yapadera, yomwe imagwira ntchito pazida zambiri za qPCR.Sikofunikira kusintha kuchuluka kwa ROX pazida zosiyanasiyana.Ndikofunikira kuwonjezera zoyambira ndi ma templates kuti mukonzekere kachitidwe kakukulitsa.

Zigawo

Universal Blue qPCR Master Mix

Zosungirako

Mankhwalawa amatumizidwa ndi mapaketi a ayezi ndipo amatha kusungidwa ku -25 ℃ ~ -15 ℃ kwa miyezi 18.M'pofunika kupewa kuwala kowala pamene mukusunga kapena kukonzekera njira yochitira.

Kufotokozera

Malangizo

1.Reaction System

[Zindikirani]: Sakanizani bwino musanagwiritse ntchito kuti mupewe thovu lochulukirapo kuti musagwedezeke mwamphamvu.

a) Kuphatikizika koyambira: Kukhazikika komaliza koyambira ndi 0.2μmol/L, ndipo kumathanso kusinthidwa pakati pa 0.1 ndi 1.0μmol/L momwe kuli koyenera.

b) Template concentration: Ngati template ndi undiluted cDNA stock solution, voliyumu yogwiritsidwa ntchito sayenera kupitirira 1/10 ya voliyumu yonse ya qPCR reaction.

c) Dilution ya template: Ndikofunikira kuti muchepetse yankho la cDNA stock ndi nthawi 5-10.Kuchuluka koyenera kwa template yowonjezeredwa ndikwabwinoko pamene mtengo wa Ct wopezedwa ndi kukulitsa ndi 20-30 mizungu.

d) Dongosolo lamachitidwe: Ndikofunikira kugwiritsa ntchito 20μL kapena 50μL kuti muwonetsetse kuchita bwino komanso kubwereza kwa kukulitsa kwa jini.

e) Kukonzekera kwadongosolo: Chonde konzekerani mu benchi yoyera kwambiri ndikugwiritsa ntchito nsonga ndi machubu ochitira popanda zotsalira za nyukiliya;tikulimbikitsidwa kugwiritsa ntchito nsonga ndi makatiriji fyuluta.Pewani kuipitsidwa ndi aerosol ndi kuipitsidwa.

2.Pulogalamu yamachitidwe

Pulogalamu Yokhazikika

Pulogalamu Yofulumira

[Zindikirani]: Pulogalamu yofulumira ndiyoyenera majini ambiri, ndipo mapulogalamu okhazikika amatha kuyesedwa pama jini amtundu wachiwiri.

a) Kutentha kowonjezera ndi nthawi: Chonde sinthani molingana ndi kutalika kwa primer ndi jini ya chandamale.

b) Kupeza siginecha ya Fluorescence (★): Chonde ikani njira yoyesera molingana ndi zofunikira mu malangizo ogwiritsira ntchito chida.

c) Mpiringidzo wosungunuka: Pulogalamu yosasinthika ya chida itha kugwiritsidwa ntchito nthawi zonse.

3. Kusanthula Zotsatira 

Zofananira zosachepera zitatu zachilengedwe zidafunikira pakuyesa kachulukidwe.Pambuyo pazimenezi, mapindikidwe a amplification ndi ma curve osungunuka ayenera kutsimikiziridwa.

3.1 Kukulitsa curve:

Mulingo wokhazikika wokulitsa ndi mawonekedwe a S.Kusanthula kachulukidwe ndikolondola kwambiri pamene mtengo wa Ct ukutsika pakati pa 20 ndi 30. Ngati mtengo wa Ct ndi wochepera 10, m'pofunika kuchepetsa template ndikuyesanso.Pamene mtengo wa Ct uli pakati pa 30-35, ndikofunikira kuonjezera tsatanetsatane wa template kapena kuchuluka kwa machitidwe, kuti mupititse patsogolo kukulitsa bwino ndikuwonetsetsa kulondola kwa kusanthula kwa zotsatira.Pamene mtengo wa Ct ndi waukulu kuposa 35, zotsatira za mayesero sizingathe kusanthula mozama mawu a jini, koma zingagwiritsidwe ntchito pofufuza za khalidwe.

3.2 Kusungunuka kopindika:

Chiwongoladzanja chimodzi chokha chachitsulo chosungunuka chimasonyeza kuti momwe zimachitikira ndi zabwino ndipo kusanthula kachulukidwe kungathe kuchitidwa;ngati nsonga yosungunuka ikuwonetsa nsonga ziwiri kapena zingapo, kusanthula kwachulukidwe sikungachitike.Mapiritsi osungunuka amawonetsa nsonga ziwiri, ndipo m'pofunika kuweruza ngati nsonga yomwe siinapangidwe ndi primer dimer kapena kukwezedwa kosagwirizana ndi DNA agarose gel electrophoresis.Ngati ndi primer dimer, tikulimbikitsidwa kuti muchepetse ndende ya primer kapena kukonzanso zoyambira ndi zokulitsa kwambiri.Ngati sikuli kukulitsa kwinakwake, chonde onjezerani kutentha kwa annealing, kapena panganinso zoyambira motsata ndondomeko.

Zolemba

Chonde valani PPE yofunikira, malaya a labu ndi magolovesi, kuti muwonetsetse thanzi lanu ndi chitetezo!

Izi ndizogwiritsidwa ntchito pofufuza POKHA!