RT-LAMP Colormetric Master Mix HCB5204A
Ngwaahịa a nwere ihe nchekwa mmeghachi omume, RT-Enzymes Mix (Bst DNA polymerase na reverse transcriptase okpomọkụ na-eguzogide), ihe nchekwa lyophilized na ihe ndị na-esiji chromogenic.Iji jiri, naanị jiri ihe nchekwa, a na-agwakọta enzyme mmeghachi omume na primer ma tinye ya na ndebiri;ịgbakwụnye ihe nchekwa lyophilized nwere ike kwụ ọtọ.Ejikọtara ya na lyophilizer na lyophilized, na naanị primers na ndebiri ka agbakwunyere mgbe ejiri ya.Ngwa a na-enye ngwa ngwa, nchọpụta anya doro anya nke mmụba, nke mmeghachi omume na-adịghị mma na-egosi na-acha uhie uhie na mmeghachi omume dị mma na-egosipụta site na mgbanwe na-acha odo odo.
Akụkụ
| Akụkụ | HCB5204A-01 | HCB5204A-02 | HCB5204A-03 |
| Ihe nchekwa Amplification agbazitere loop (ya na agba) | 0.96 ml | 4.80 ml × 2 | 9.60 ml × 10 |
| Ngwakọta RT-enzymes | 270 μl | 2.70 ml | 2.70 ml × 10 |
| Lyophilized nchebe | 0.96 ml × 2 | 9.60 ml × 2 | 9.60 ml × 20 |
Ngwa
Maka DNA ma ọ bụ RNA isothermal amplification.
Ọnọdụ Nchekwa
Ejiri akpụrụ akpụ ebu, echekwara na -25 ~ -15 ℃.Zere na-agbaze oyi mgbe niile, ngwaahịa a dị irè maka ọnwa 12.
Protocol
1.Kpee ihe nchekwa mmeghachi omume a ga-eji na ọnụ ụlọ okpomọkụ.Vortex dị nkenke ma ọ bụ tụgharịa tubes ọtụtụ oge ka ị gwakọta nke ọma, wee centrifuge ịnakọta mmiri mmiri na ala nke tube ahụ.
2.Nkwadebe nke usoro mmeghachi omume.Enwere ike ịkwadebe reagent a na sistemụ mmeghachi omume abụọ, ngwakọta mmeghachi omume mmiri mmiri na ngwakọta sistemu lyophilized.
1) Kwadebe ngwakọta mmeghachi omume mmiri mmiri
| Akụkụ | Olu |
| Ihe nchekwa Amplification agbazitere loop (ya na agba) | 10 μl |
| Ngwakọta RT-enzymes | 2.8 μl |
| 10 × Ngwakọta mbụa | 5 μl |
| Ụdị DNA/RNA b | × μL |
| Mmiri na-enweghị nuklia | Ihe dị ka 50 μl |
2) Lyophilization usoro mix
① Kwadebe ngwakọta lyophilized
| Akụkụ | Olu |
| Ihe nchekwa Amplification agbazitere loop (ya na agba) | 10 μl |
| Lyophilized nchebe | 20 μl |
| Ngwakọta RT-enzymes | 2.8 μl |
| Mmiri na-enweghị nuklia | Ihe dị ka 50 μl |
② Lyophilization: Ngwakọta a kwadebere bụ lyophilized na usoro 50μL
③ Kwadebe ngwakọta mmeghachi omume
| Akụkụ | Olu |
| Ngwakọta Lyophilized | 1 ibe |
| 10 × Ngwakọta mbụa | 5 μl |
| Ụdị DNA/RNA b | × μL |
| Mmiri na-enweghị nuklia | Ihe dị ka 50 μl |
ndetu:
1) a.10 × Primer Mix : 16 μM FIP / BIP, 2 μM F3 / B3, 4 μM Loop F / B;
2) b.DEPC (mmiri soluble) ka akwadoro maka templ acid nucleic.
1.Tinye na 65 Celsius C maka 30-45mins, nke enwere ike ịgbatị nke ọma dịka oge mgbanwe agba si dị.
2.Dị ka anya gba ọtọ si kwuo, odo na-adị mma na uhie dị njọ.
Ihe ndetu
1.Nnu nwere ike ịpụta na ala nke tube nchekwa, vortex obere oge ma ọ bụ tụgharịa tubes ọtụtụ oge iji gwakọta nke ọma na ụlọ okpomọkụ.
2.Enwere ike ịhazi okpomọkụ mmeghachi omume n'etiti 62 ℃ na 68 ℃ dị ka ọnọdụ nke primers si dị.
3.The ngwugwu reagents ekwesịghị ikpughe na ikuku ruo ogologo oge.
4.Mmeghachi omume ngbanwe nke uhie na odo na-adabere na pH mgbanwe nke sistemu mmeghachi omume, biko ejila ihe nchekwa nchekwa Tris nucleic acid nwere, akwadoro iji ddH.2O echekwabara nucleic acid;
5.Ekwesịrị ịhazi nnwale ahụ, gụnyere nkwadebe nke usoro mmeghachi omume, lyophilization, na nhazi nlele na usoro ntinye ihe nlele;
6.Iji zere mmetọ, a na-atụ aro ka ịkwadebe usoro mmeghachi omume na bench dị ọcha, na ndị ọzọ Tinye ndebiri na mkpuchi uzuzu nke ụlọ ahụ iji zere nnyonye anya ụgha.
