Escherichia Coli O157: H7 Nucleic Acid Detection Kit (PCR Fluorescent Probe Method/Lyophilization)
ʻO ka wehewehe ʻana o ka pahu:
Cat.No.FP103
Hoʻohana ʻia ia no ka ʻike wikiwiki ʻana a me ka nānā ʻana o E. coli O157:H7 i ka meaʻai, nā meaʻai, nā ʻano wai a me nā laʻana kaiapuni.
Nā wehewehe
Hoʻohana ʻia noʻike wikiwiki a me ka nānā ʻana o E. coli O157:H7 i ka meaʻai, nā meaʻai, nā laʻana wai a me nā laʻana kaiapuni.
[Kumu hoʻāʻo]
Wahi a ke kumu o ka ʻenehana PCR fluorescent, ua hoʻolālā ʻia nā primers kikoʻī a me Taqman probes no ka gene kikoʻī o Escherichia coli O157: H7, a ʻike ʻia e kahi mea hana PCR fluorescent, no laila e ʻike i ka ʻike ʻana o Escherichia coli O157: H7 Qualitative detection of DNA.
Kiko kiko
'Ōlelo Aʻo: ʻAʻole i hoʻokomo ʻia ka probe channel ROX.
| Cnā mea hoʻokūkū | Hōʻike | Quantity |
| Hoʻopaʻa A | Paipu | 1 |
| Hoʻopaʻa B | Paipu | 1 |
| Manaʻo maikaʻi | Paipu | 1 |
| Manaʻo ʻino | Paipu | 1 |
Hoʻohana Manaʻo ʻia
Hoʻohana ʻia no ʻike wikiwiki a me ka nānā ʻana o E. coli O157:H7 i ka meaʻai, nā meaʻai, nā laʻana wai a me nā laʻana kaiapuni.
Nā Kūlana Waihona a me ka lā pau
E mālama i ka -20 ℃ i ka pōʻeleʻele a pale i ka hau a me ka hoʻoheheʻe pinepine ʻana.
ʻO 12 ka manawa kūponomahina, a hōʻike ʻia ka lā hana ma ka ʻeke o waho.
Na Mea Hana A me na Mea Hoohana
ʻO ka mea hana PCR ka nui o ka fluorescent, ka pū pipette a me nā ʻōlelo aʻoaʻo like, vortex shaker, mini centrifuge.
Hoʻohana
1. Laʻana Hana
1.1 ʻAno laʻana: He kūpono kēia pahu no ka meaʻai, nā meaʻai, nā ʻano wai a me nā mea hoʻohālike ʻē aʻe i manaʻo ʻia e hoʻohaumia ʻia e Escherichia coli O157:H7.No nā huahana ʻiʻo i hoʻoponopono hohonu ʻia, nā mea inu a me nā mea ʻē aʻe i loaʻa nā puaʻa, pono lākou e holoi ʻia e pale aku i ka hopena o ka hōʻiliʻili hōʻailona fluorescence.
1.2 Laʻana hana: E nānā i "GB 4789.10-2016 Food Safety National Standard Food Microbiological Examination o Escherichia coli O157: H7 Test" no ka laʻana hoʻomākaukau, enrichment moʻomeheu a me ka kaawale o Escherichia coli O157: H7.
- Nka ʻili ʻana o ka ʻakika ucleic
E lawe i 20 mL o ka wai hoʻonui i loko o ka 1.5 mL centrifuge tube, e hoʻohui i 200 μL o ka microbial lysate (pono ʻia ka pahu hou), vortex no 30 kekona, centrifuge pōkole, a waiho ʻia.
Manaʻo: Pono e hoʻopau ʻia ka unuhi ʻana o ka waikawa nucleic mai ka lysate i loko o 10 mau minuke, ʻaʻole hiki ke mālama ʻia no ka manawa lōʻihi.
3. Hoʻonui ʻia ka ʻakika Nucleic
3.1 E ho'ā i ka mea hana PCR quantitative fluorescent no ka hoʻohana.
Buffer A a me Buffer B mai ka pahu, hoʻoheheʻe maikaʻi iā lākou, a centrifuge pōkole.E hoʻohui i 18 μL Buffer A a me 2 μL Buffer B i kēlā me kēia paipu hoʻoheheʻe PCR.A laila e hoʻohui i 5 mL i kēlā me kēia o ka mana ʻino, ka wai nucleic i unuhi ʻia, a me ka mana maikaʻi i loko o nā paipu hopena PCR, e uhi i nā paipu, a me ka centrifuge pōkole.
3.3 E hoʻololi i ka paipu hoʻoheheʻe PCR i ka mīkini PCR fluorescent, a e hoʻohana i nā kaʻina hana e hana ai i nā hoʻokolohua hoʻonui: koho i 25 mL no ka ʻōnaehana pane, hōʻiliʻili i nā hōʻailona fluorescence ma 60°C no kēlā me kēia pōʻai, a koho i ka FAM no ke ala ʻike.
| ʻanuʻu | Papahana | Ka helu o nā pōʻaiapuni |
| 1 | 37℃ 5min | 1 |
| 2 | 9 5 ℃ 3min | 1 |
| 3 | 95°C 15s | 4 0 |
| 60 ℃ 30s (hōʻiliʻili fluorescence) |
Nā alakaʻi no ka nānā ʻana i nā pilikia
The following is an analysis of the problems that might be encountered in the extraction of viral RNA. We wish it would be helpful to your experiment. In addition, for other experimental or technical problems other than operating instructions and problem analysis, we have dedicated technical support to help you. Contact us if you need at : 028-83361257or E-mail：Tech@foregene.com。
ʻAʻole hiki ke unuhi ʻia ka RNA a i ʻole he haʻahaʻa ka hua o ka nucleic acid
Nui nā mea e pili ana i ka hoʻihoʻi ʻana, e like me: sample RNA content, ala o ka hana, elution volume, etc.
Ka nānā 'ana i nā kumu ma'amau:
1.Ice bath a haʻahaʻa-mehana (4 ° C) centrifugation i ka wā hana.
Manaʻo: Hana wela lumi (15-25 ° C), ʻaʻole ʻauʻau hau a me ka centrifuge wela haʻahaʻa.
2. Ka mālama pono ʻole ʻana i ka laʻana a i ʻole ka waiho ʻana o ka laʻana no ka lōʻihi loa.
Manaʻo Manaʻo: E kūʻai i nā laʻana ma -80 ° C a i ʻole i loko o ka hau hau i loko o ka naikokene wai, a pale i ka hoʻohana pinepine ʻana i ka hau hau;e ho'āʻo e hoʻohana i nā laʻana i hōʻiliʻili hou ʻia no ka unuhi ʻana o RNA.
3.Insufficient hāpana lysis
Manaʻo Manaʻo: E ʻoluʻolu e ʻoluʻolu e hui pū ʻia ka hāpana a me ka hopena hana (Linear Acrylamide) a hoʻomoʻi ʻia no 10 min ma ka lumi wela (15-25 ° C)
4. Ua hoʻohui hewa ʻia ka eluent
Manaʻo Manaʻo: E hōʻoia e hoʻohui ʻia ka RNase-Free ddH2O i ka waena o ka membrane o ke kolamu hoʻomaʻemaʻe.
5. Improper leo o anhydrous ethanol ma Buffer viRW2
Manaʻo Manaʻo: E ʻoluʻolu e hahai i nā ʻōlelo aʻoaʻo, e hoʻohui i ka nui kūpono o ka ethanol anhydrous i Buffer viRW2 a hui maikaʻi ma mua o ka hoʻohana ʻana i ka pahu.
6.Improper laʻana hoʻohana.
Manaʻo: 200µl o ka laʻana no 500μl o Buffer viRL.ʻO ka nui o ka nui o ka laʻana e hoʻemi ʻia ka RNA extraction rate.
7. Improper elution volume a i ʻole elution piha ʻole.
Manaʻo: ʻO ka leo eluent o ke kolamu hoʻomaʻemaʻe ʻo 30-50μl;inā ʻaʻole ʻoluʻolu ka hopena elution, pono e hoʻohui i ka RNase-Free ddH pre-heated.2O a hoʻolōʻihi i ka manawa kau ma ka lumi wela, e like me 5-10min
8. He koena ethanol ke kolamu hoʻomaʻemaʻe ma hope o ka holoi ʻana i Buffer viRW2.
Manaʻo Manaʻo: Inā e mau ana ka ethanol ma hope o ka holoi ʻana i Buffer viRW2 a me ka centrifugation pahu pahu ʻole no 2min, hiki ke waiho ʻia ke kolamu hoʻomaʻemaʻe ma ke ana wela no 5min ma hope o ka centrifugation pahu-wai ʻole e wehe loa i ke koena ethanol.
ʻO ka hoʻohaʻahaʻa ʻana o nā molekala RNA i hoʻomaʻemaʻe ʻia
ʻO ka maikaʻi o ka RNA i hoʻomaʻemaʻe ʻia e pili ana i nā mea e like me ka waiho ʻana o ka laʻana, ka haumia RNase, a me ka hana.
Ka nānā 'ana i nā kumu ma'amau:
1. ʻAʻole mālama ʻia nā laʻana i hōʻiliʻili ʻia i ka manawa.
Manaʻo Manaʻo: Inā ʻaʻole hoʻohana ʻia ka hāpana i ka manawa ma hope o ka hōʻiliʻili ʻana, e ʻoluʻolu e mālama iā ia ma -80 ℃ a i ʻole ka nitrogen wai koke.No ka unuhi ʻana i nā molekole RNA, e hoʻāʻo e hoʻohana i nā laʻana i hōʻiliʻili hou ʻia inā hiki.
2. ʻO nā laʻana i hōʻiliʻili ʻia i ka hau a hoʻoheheʻe pinepine ʻia.
Manaʻo Manaʻo: E pale i ka maloʻo a me ka hoʻoheheʻe pinepine ʻana (ʻaʻole i ʻoi aku ma mua o hoʻokahi manawa) i ka wā o ka hōʻiliʻili ʻana a me ka mālama ʻana, inā ʻaʻole e emi ka hua o ka waika nucleic.
3. Ua hoʻokomo ʻia ʻo RNase i loko o ka lumi hana a ʻaʻole i hoʻohana ʻia nā mīkina lima, nā maka, a me nā mea ʻē aʻe.
Manaʻo Manaʻo: E hana maikaʻi ʻia ka hoʻāʻo ʻana o nā moleke RNA i loko o kahi lumi hana RNA ʻokoʻa, a hoʻomaʻemaʻe ʻia ka papa hoʻokolohua ma mua o ka hoʻokolohua.E hoʻohana i nā mīkina lima a me nā maka i ka wā o ka hoʻokolohua e pale aku i ka hōʻino ʻana o RNA ma muli o ka hoʻokomo ʻana o RNase.
4. Ua haumia ka reagent e RNase i ka wā o ka hoʻohana.
Manaʻo: Hoʻololi me ka Viral RNA Isolation Kit hou no nā hoʻokolohua pili.
5. ʻO ka RNase contamination o nā paipu centrifuge, nā ʻōlelo pipette, a me nā mea ʻē aʻe.
Ua hoʻopili ʻia nā molekala RNA i hoʻomaʻemaʻe ʻia i nā hoʻokolohua o lalo
ʻO nā molekala RNA i hoʻomaʻemaʻe ʻia e ke kolamu hoʻomaʻemaʻe e hoʻopili i nā hoʻokolohua i lalo inā he nui nā ion paʻakai a i ʻole nā protein, e like me: reverse transcription, Northern Blot, etc.
1. Aia nā ion paʻakai i koe i loko o nā molekele RNA i hoʻohemo ʻia.
Manaʻo Manaʻo: E hōʻoia ua hoʻohui ʻia ka leo kūpono o ka ethanol anhydrous i Buffer viRW2, a holoi i ke kolamu hoʻomaʻemaʻe ʻelua e like me ka wikiwiki centrifugation pololei ma nā kuhikuhi hana; Inā he mau iona paʻakai i koe, hiki iā ʻoe ke hoʻohui i ka Buffer viRW2 i ke kolamu hoʻomaʻemaʻe, a waiho i ka lumi wela no 5min.A laila e hana i ka centrifugation e wehe i nā ion paʻakai i ka nui loa
2. Aia ke koena ethanol i loko o nā molekele RNA i hoʻokahe ʻia
Manaʻo Manaʻo: i ka manawa e hōʻoia ai ua holoi ʻia nā kolamu hoʻomaʻemaʻe e Buffer viRW2, e hana i ka centrifugation pahu ʻole e like me ka wikiwiki centrifugal ma nā kuhikuhi hana.Inā he ethanol koe, hiki ke waiho ʻia no 5 mau minuke ma ka lumi wela ma hope o ka hoʻoheheʻe ʻia ʻana o ka ethanol no ka wehe ʻana i ke koena ethanol i ka nui loa.
Nā Papa kuhikuhi:
