Intambwe imwe RT-qPCR SYBR Icyatsi kibisi
Injangwe No: HCB5140A
Intambwe imwe RT-qPCR Syber Green Premix ni iyo kugereranya fluorescence ishingiye kuri SYBR Icyatsi I Irangi.Ukoresheje primers yihariye ya gene, reaction transcript hamwe na qPCR reaction irangizwa mumiyoboro imwe, bivanaho gukenera inshuro nyinshi gufungura no gufungura imiyoboro, kunoza cyane imikorere no kugabanya ibyago byo kwanduza.Kuburugero rwa RNA, ibikoresho bikoresha ubushyuhe bwihanganira Reverse Transcriptase kugirango ikoreshwe neza cDNA hamwe na HotStart Taq ADN Polymerase kugirango yongere umubare.Munsi ya sisitemu ya buffer nziza, ibyiyumvo byibikoresho birashobora kuba hejuru ya 0.1 pg kubigenewe byerekanwe cyane kandi hejuru ya 1 pg kubigaragaza neza.Igikoresho gikwiranye no kongera urugero rwa ADN.Ifasha kumenya neza no kugereranya acide nucleic biva mu bimera bitandukanye by’inyamanswa n’inyamaswa, selile na mikorobe.
Ibigize
No | Izina | Umubumbe | Umubumbe |
1 | Buffer Yambere | 250 μL | 2 × 1.25 mL |
2 | Kuvanga Enzyme Yambere | 20 μL | 200 μL |
3 | RNase Ubuntu H.2O | 250 μL | 2 × 1.25 mL |
Ububiko
Ibicuruzwa bigomba kubikwa kuri -25 ~ -15 ℃ kure yumucyo kumwaka 1.
Amabwiriza
1.Iboneza rya sisitemud
Ibigize | Umubumbe (μL) | Umubumbe (μL) | Kwibanda kwanyuma |
Buffer Yambere | 12.5 | 25 | 1 × |
Kuvanga Enzyme Yambere | 1 | 2 | - |
Imbere Imbere (10 μ mol / L)a | 0.5 | 1 | 0.2 μ mol / L. |
Subiza Primer (10 μ mol / L)a | 0.5 | 1 | 0.2 μ mol / L. |
RNA Tamplateb | X | X | - |
RNase Ubuntu H.2O.c | kugeza 25 | kugeza kuri 50 | - |
Inyandiko:
1) a.T.yibanze bwa primer yibanze yari 0.2 μ mol / L, nayo ishobora guhinduka hagati ya 0.1 na 1μmol / L nkuko bikwiye.
2) b.Reagent irakomeye cyane, hamwe na RNA Yuzuye murwego rwa 1pg-1μg, kandi gupima ingero zabantu byerekanaga ko byinjijwe neza muri 1 pg-100 ng, bigenzura agaciro ka Ct muri rusange hagati ya 15-30 nkuko bikwiye.
3) c.Birasabwa gukoresha 20μL cyangwa 50μL kugirango tumenye neza kandi byororoke byintego ya gene amplification.
4) d.Nyamuneka tegura intebe ya ultra-isukuye kandi ukoreshe inama zidafite ibisigisigi bya nuclease hamwe nigituba cya reaction;inama hamwe na filteri ya karitsiye irasabwa.Irinde kwanduza umusaraba no kwanduza aerosol.
2.Gahunda yo kubyitwaramo
Intambwe | Ubushuhe. | Igihe | Amagare |
Guhindura inyandiko | 50 ℃a | 6min | 1 |
Gutandukana kwambere | 95 ℃ | 5min | 1 |
Amplification reaction | 95 ℃ | 15 amasegonda | 40 |
60 ℃b | 30 amasegonda | ||
Gushonga umurongo | Igikoresho gisanzwe | 1 |
Inyandiko:
1) a.Ubushyuhe bwo guhinduranya bushobora guhitamo hagati ya 50-55 ° C ukurikije ibikenewe mu bushakashatsi.Kuburugero rwa ADN, inzira yo kwandukura irashobora guhinduka.
2) b.Mubihe bidasanzwe ubushyuhe bwa annealing / kwaguka burashobora guhinduka ukurikije primer Tm agaciro, 60 ° C birasabwa.
Inyandiko
1. Iki gicuruzwa nikoreshwa mubushakashatsi gusa.
2. Nyamuneka kora ikoti ya laboratoire hamwe na gants imwe ikoreshwa, kubwumutekano wawe.