Kit ɗin qPCR Ta Wayar salula Kai tsaye—SYBR GREEN I Direct Cell Lysis Cell Shirye-shiryen qRT-PCR Kits-mataki ɗaya
Bayani
Wannan kit ɗin yana amfani da tsarin buffer na musamman wanda zai iya sakin RNA da sauri daga samfuran ƙwayoyin halitta don halayen RT-qPCR, ta haka ne ke kawar da tsarin tsarkakewar RNA mai cin lokaci da wahala.Ana iya samun samfurin RNA a cikin mintuna 7 kawai.Na 5×Direct RT Mix da 2×Kai tsaye qPCR Mix-SYBR reagents wanda kit ɗin ya samar zai iya samun sauri da inganci don samun sakamako na PCR na ainihin lokaci.
5×Direct RT Mix da 2×Direct qPCR Mix-SYBR suna da ƙarfin juriya mai hanawa, kuma ana iya amfani da lysate na samfuran azaman samfuri don RT-qPCR kai tsaye.Wannan kit ɗin ya ƙunshi keɓaɓɓen RNA high-affinity Foregene reverse transcriptase, da Hot D-Taq DNA polymerase, dNTPs, MgCl.2, amsawa buffer, PCR ingantawa da stabilizer.
Ƙayyadaddun bayanai
200×20μl Rxns, 1000×20μl Rxn
Abubuwan Kit
| Kashi na I | Buffer CL |
| Foregene Protease Plus II | |
| Buffer ST | |
| Kashi na II | Goge DNA |
| 5× Direct RT Mix | |
| 2× Direct qPCR Mix-SYBR | |
| 50× ROX Reference Dye | |
| RNase-Free ddH2O | |
| Umarni | |
Fasaloli & fa'idodi
■Mai sauƙi da inganci: tare da fasahar Cell Direct RT, ana iya samun samfuran RNA a cikin mintuna 7 kawai.
∎ Bukatar samfurin karami ne, kamar yadda za a iya gwada ƙananan sel guda 10.
∎ Babban abin da ake fitarwa: zai iya gano RNA da sauri a cikin sel waɗanda aka tsara a cikin faranti 384, 96, 24, 12, 6-rijiya.
∎ Gogewar DNA na iya saurin cire kwayoyin halittar da aka fitar, da rage tasirin sakamakon gwaji na gaba.
∎ Ingantaccen tsarin RT da qPCR yana sa jujjuyawar matakai biyu na RT-PCR mafi inganci kuma PCR ta fi ƙayyadaddun ƙayyadaddun bayanai, kuma mafi juriya ga masu hana amsawar RT-qPCR.
Kit aikace-aikace
Iyakar aikace-aikace: sel masu al'ada.
- RNA da aka saki ta samfurin lysis: kawai ana amfani da samfurin RT-qPCR na wannan kit.
- Za a iya amfani da kit ɗin don dalilai masu zuwa: nazarin maganganun kwayoyin halitta, tabbatar da tasirin sirna mai tsaka-tsaki na yin shiru, gwajin magunguna, da sauransu.
zane
Adana da Rayuwar Shelf
Ya kamata a adana sashe na I na wannan kit a 4 ℃;Ya kamata a adana Sashe na II a -20 ℃.
Ya kamata a adana Foregene Protease Plus II a 4℃, kar a daskare a -20 ℃.
Reagent 2×Ya kamata a adana qPCR Mix-SYBR kai tsaye a -20℃a cikin duhu;idan ana amfani dashi akai-akai, ana iya adana shi a 4℃ don ajiya na ɗan gajeren lokaci (amfani da sama a cikin kwanaki 10).
Ka'idodin ƙira na ainihin lokacin PCR
Gaba Farawa da Juya Farko
Don Real Time PCR, ƙirar farko tana da mahimmanci.Abubuwan haɓaka suna da alaƙa da ƙayyadaddun ƙayyadaddun ƙayyadaddun ƙimar PCR, kuma ana iya tsara su tare da la'akari da ƙa'idodi masu zuwa:
Tsayin farko: 18-30bp.
GC abun ciki: 40-60%.
Ƙimar Tm: Software na ƙira na farko, kamar Primer 5, na iya ba da ƙimar Tm na firamare.Ma'auni na Tm na sama da na ƙasa ya kamata su kasance kusa da iyawa.Hakanan ana iya amfani da dabarar lissafin Tm: Tm = 4 °C (G + C) + 2 °C (A + T).Lokacin yin PCR, ana zaɓar zafin da ke ƙasa da ƙimar ƙimar Tm na 5 ° C gabaɗaya azaman zazzabi mai ɗaukar nauyi (madaidaicin haɓakar zafin jiki na iya ƙara ƙayyadaddun halayen PCR).
Alamar farko da samfuran PCR:
Tsawon samfurin PCR na ƙirar ƙira ya fi dacewa 100-150bp.
Ya kamata a kauce wa ƙera ƙirar ƙira a cikin yanki na biyu na samfuri gwargwadon yiwuwa.
Guji samuwar tushe guda 2 ko sama da haka tsakanin 3′ na ƙofofin sama da na ƙasa.
Babban tushe 3′ na tashar ba zai iya kasancewa tare da ƙarin ƙarin 3 a jere G ko C.
Masu farawa da kansu ba za su iya samun ƙarin tsari ba, in ba haka ba za a kafa tsarin gashin gashi, yana shafar haɓaka PCR.
Yakamata a rarraba ATCG daidai gwargwado kamar yadda zai yiwu a cikin jeri na farko, kuma ya kamata a guje wa tushe na 3′ a matsayin T.
Shafi 1: Tantanin halitta Direct RT-qPCR Kit na ƙarin fakitin
1.Cell Lysis Magani
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| Abubuwan Kit (24-riji tsarin lysis / rijiyar) | Saukewa: DRT-01011-A1 | Saukewa: DRT-01011-A2 | |
| 100 T | 500 T | ||
| SasheI | Buffer CL | 20 ml | 100 ml |
| Foregene Protease Plus II | 400 ml | 1 ml × 2 | |
| Buffer ST | 1 ml × 2 | 10 ml | |
| SasheII | Goge DNA | 400 ml | 1 ml × 2 |
2.RT Mix
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| Abubuwan Kit (20 μl tsarin amsawa) | Saukewa: DRT-01011-B1 |
| 200 T | |
| 5× Direct RT Mix | 800 ml |
| RNase-Free ddH2O | 1.7 ml × 2 |
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| Abubuwan Kit (20 μl tsarin amsawa) | Saukewa: DRT-01011-C1 | Saukewa: DRT-01011-C2 |
| 200 T | 1000 T | |
| 2× Direct qPCR Mix-SYBR | 1 ml × 2 | 1.7 ml × 6 |
| 50× ROX Reference Dye | 40ml ku | 200 μl |
| RNase-Free ddH2O | 1.7 ml | 10 ml |
Jagoran Jagora:
