New Coronavirus(SARS-Cov-2) Kete Rapu Waikawa Nucleic
New Coronavirus(SARS-Cov-2) Nucleic Acid Detection Kit
(Fluorescent RT-PCR Probe Method) Product Manual
【Product name 】Coronavirus Hou(SARS-Cov-2) Kete Tirohanga Waikawa Nucleic (Tikanga Tirohanga RT-PCR Fluorescent)
【Packaging specifications 】25 Whakamātautau/Ket
【Intended ustau】
Ka whakamahia tenei kete mo te rapunga tohu o te waikawa karihi mai i te coronavirus hou i roto i nga ruuma nasopharyngeal, oropharyngeal (korokoro) swabs ihu o mua, swips mid-turbinate, horoi ihu me te nasal aspirates mai i nga tangata e whakapaetia ana he COVID19 e to ratou kaiwhakarato hauora. Ko te kitenga o nga ira ORF1ab me te N o te coronavirus hou ka taea te whakamahi mo te taatai awhina me te aro turuki i nga mate coronavirus hou.
【Principles of the procedure 】
I hoahoatia tenei kete mo nga rangahau TaqMan motuhake me nga kaupapa tuatahi i hangaia mo te pukapuka coronavirus (SARS-Cov-2) ORF1ab me nga raupapa ira N. Kei roto i te otinga tauhohenga PCR e 3 nga huinga tuatahi me nga rangahau rewharewha mo te rapunga motuhake o nga whaainga, a ka whakamahia he huinga taapiri o nga tohu tuatahi me nga rangahau kareoro hei whakahaere paerewa o roto o te kete mo te kite i nga ira tiaki whare.
Ko te maataapono whakamatautau ko te mea ka kerihia, ka whakahekehia e te mahi exonuclease o te Taq enzyme i roto i te tauhohenga PCR, kia wehea ai te roopu rekoata korero me te roopu rewharewha kua tineia, kia taea ai e te punaha aro turuki i te rewharewha. tohu, katahi Na roto i te whakarangatiratanga o te whakakaha PCR, ka tae te tohu orooro o te rangahau ki te uara paepae kua whakaritea-Ct uara (Tauwhiu Paepae). Mena karekau he amplicon whainga, ka tata te roopu kairipoata o te rangahau ki te roopu tiini. I tenei wa, ka puta te whakawhitinga o te kaha o te resonance resonance, a, ka tineia te rere o te roopu kairipoata e te roopu tinei, kia kore ai e kitea te tohu o te hiko e te taputapu PCR fluorescent.
Hei aro turuki i te whakamahinga o nga reagents i te wa o te whakamatautau, kua rite te kete ki nga mana pai me te kino: kei roto i te mana pai te plasmid recombinant pae ūnga, me te mana kino ko te wai Maturu, e whakamahia ana ki te aro turuki i te parahanga taiao. E taunaki ana kia whakatauhia he mana pai me te mana kino i te wa e whakamatautau ana.
【Main components 】
| Cat. No. | BST-SARS-25 | BST-SARS-DR-25 | Huingaents | |
| Name | Specwhakau | Te mahatenei | Te mahatenei | |
| Te whakahaere pai | 180 μL / ipu | 1 | 1 | Ko nga plasmids i hangaia he mea hanga, Te wai maturu |
| Mana kino | 180 μL / ipu | 1 | 1 | Te wai marewa |
| SARS-Cov-2 Mix | 358.5 μL/ ipu | 1 | / | Nga takirua tuatahi, te rapunga maataki karewharewha motuhake, nga dNTP, , MgCl2, KCl, Tris-Hcl, Wai Maturu, aha atu |
| Whakaranu Enzyme | 16.5 μL/ ipu | 1 | / | Taq enzymes, reverse transcriptase, UNG enzymes, etc. |
| SARS-Cov-2 Mix(Lyophilised) | 25 nga whakamatautau / ipu | / | 1 | Ko nga takirua tuatahi, ko nga maataki karewharewha whakakitenga motuhake, dNTPs, Taq enzymes, whakamuri transcriptase, Wai Maturu, aha atu. |
| 2x Kaipupuri | 375 μL/pēpu | / | 1 | MgCl2, KCl, Tris-Hcl, Wai Maturu, etc. |
Tuhipoka：(1) Ko nga waahanga kei roto i nga kete puranga rereke kaore e taea te whakauru, te whakawhiti ranei.
(2) Whakaritea taau ake reagent: kete tangohanga waikawa nucleic.
【Storage conditions a expiration date 】
For BST-SARS-25:Te kawe me te rokiroki i te -20±5 ℃ mo te wa roa.
For BST-SARS-DR-25:Te kawe i te pāmahana rūma. Rokiroki ki te -20±5 ℃ mo te wa roa.
A ape i nga huringa whakatio-rewa. Ko te waa whaimana kua whakaritea mo te 12 marama.
Tirohia te tapanga mo te ra i hangaia me te whakamahi.
I muri i te whakatuwheratanga tuatahi, ka taea te penapena i te reagent ki te -20±5 ° C mo te kore neke atu i te 1 marama, tae noa ki te mutunga o te wa reagent, ahakoa te ra ka tae tuatahi mai, hei karo i nga huringa whakatio-rewa, me te maha o te reagent tio. -kaua e neke ake i te 6 nga wa o nga huringa whakamaroke.
【Applicable instrument】ABI 7500, SLAN-96P, Roche-LightCycler-480.
【Sample requirements 】
1.Momo tauira e tika ana: Te otinga waikawa karihi kua tangohia.
2.Tauira rokiroki me te kawe waka: Rokiroki i-20±5 ℃ mo nga marama 6. Whakatio me te whakamaroke i nga tauira kaua e neke atu i te 6 nga wa.
【Testing method】
1.Nucleic acid extraction
Tīpakohia he kete tangohanga waikawa karihi e tika ana hei tango i te waikawa nucleic viral, ka whai i nga tohutohu kete. E taunaki ana kia whakamahia he kete tangohanga waikawa Nucleic me te kete purenga i hangaia e Yixin Bio-Tech (Guangzhou) Co., Ltd., he kete purenga waikawa karihi rite ranei.
2. Reaction reagent preparation
2.1 For BST-SARS-25:
( 1) Tango i te SARS-Cov-2 Mix and Enzyme Mix , ka rewa rawa i te pāmahana rūma uru rawa ki te taputapu Vortex katahi ka whakahiahia poto.
(2) 16.5uL Enzyme Mix i tapirihia ki te 358.5uL SARS-Cov-2 Mix ka tino whakaranu kia whiwhi i te otinga tauhohenga whakauru.
(3) Whakapaia he ngongo PCR 0.2 mL ma, ka tohu ki te 15uL o te wairewa tauhohenga whakauru i runga ake nei mo ia poka.
(4) Taapirihia te 15 μL o te wairewa waikawa karihi kua purea, te mana pai me te mana kino, ka ata kapi i te potae ngongo octal.
(5) Whakaranu pai ma te huri whakahurihuri ki raro, ka tere te centrifuge kia kuhu te wai ki raro o te ngongo.
1
2.2 For BST-SARS-DR-25:
( 1) Taapirihia te 375ul 2x Buffer ki SARS-Cov-2 Mix((Lyophilised) ki te whakarite i te ranunga tauhohenga. Whakaranu paitia ma te paipa, katahi ka centrifuge poto. （Ka rite te ranunga tauhohenga, ka tūtohu kia rongoa ki te -20℃ i rokiroki mo te wa roa.)
(2) Whakapaia he ngongo 0.2 mL PCR octal ma ka tohua ki te 15μL o te ranunga tauhohenga mo ia puna.
(3) Taapirihia te 15μL o te otinga waikawa karihi kua purea, te mana pai me te mana kino, ka ata kapi i te potae ngongo octal.
(4) Whakaranu paitia ma te huri whakamuri, ka tere te centrifuge kia kuhu te wai ki raro o te ngongo.
3. PCR amplification （Tena tirohia te pukapuka taputapu mo nga tautuhinga mahi.）
3. 1 Whakanohoia te PCR 8-tube ki roto i te ruma tauira o te taputapu PCR fluorescent, ka tautuhi i te tauira hei whakamatautau, te mana pai me te mana kino kia rite ki te raupapa o te utaina.
3.2 Te hongere rapu korikori:
( 1) Ko te ira ORF1ab te whiriwhiri i te hongere rapu o FAM (Kaipoata: FAM, Quencher: Kore).
(2) Ka tohua e te ira N te hongere kitenga o VIC (Kaipoata: VIC, Quencher: Kore).
(3) Ka tohua e te ira paerewa o roto te hongere kitenga o CY5 (Kaipoata: CY5, Quencher: Kore).
(4) Kua whakaritea te Tohutoro Passive ki ROX.
3.3 Tautuhinga tawhā papatono PCR:
| Hipanga | Te pāmahana(℃) | Te wa | Te maha o nga huringa | |
| 1 | Tauhohenga whakawhiti whakamuri | 50 | 15 min | 1 |
| 2 | Taq enzyme whakahohenga | 95 | 2.5 min | 1 |
| 3 | Taq enzyme whakahohenga | 93 | 10 s | 43 |
| Ko te toronga whakaene me te tango i te korikori | 55 | 30 s | ||
I muri i te tautuhi, tiakina te konae ka whakahaere i te hotaka tauhohenga..
4.Results analysis
Ka mutu te hotaka, ka tiakina aunoatia nga hua, ka tātarihia te pihi whakakaha. Kua whakaritea te ānau whakakaha ki te paepae taunoa taputapu.
【Explanation of test results 】
1. Whakatauhia te mana o te whakamatautau: Ko te mana pai FAM, te hongere VIC me whai anau whakakaha angamaheni, me te uara Ct i te nuinga o te waa he iti iho i te 34, engari ka rereke pea na te rereke o nga tautuhinga paepae o nga taputapu rereke. Ko te mana kino FAM, te hongere VIC kia kore e whakakaha Ct. E whakaaehia ana me tutuki nga whakaritenga i runga ake nei i te wa ano, ki te kore ka muhu tenei whakamatautau.
2. Whakawa whakatau
| FAM/VIC hongere | Te hua whakawa |
| Ct＜37 | He pai te whakamatautau tauira |
| 37≤Ct＜40 | He S-ahua te ānau whakakaha, a me tirotiro ano nga tauira whakapae; ki te rite tonu nga hua o te tirotiro ano, ka whakatauhia he pai, ki te kore he kino |
| Ct≥40 Kaore ranei he Whakanui | He kino te whakamatautau tauira (i raro ranei i te tepe o raro o te kitenga kete) |
Tuhipoka: ( 1) Mena he pai te hongere FAM me te hongere VIC i te wa kotahi, ka whakatauhia te SARS-Cov-2 kia pai.
(2) Mena he pai te hongere FAM, te hongere VIC ranei, he kino tetahi atu hongere, me whakahoki ano te whakamatautau. Mena he pai i te wa ano, ka whakatauhia ko te SARS-Cov-2 pai, ki te kore ka whakatauhia ko te SARS-Cov-2 kino.
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