Nasihu don Inganta Farfaɗowar Manna

1. Ƙara nauyin samfurin a lokacin electrophoresis.

2. Yi amfani da buffer electrophoresis da aka shirya.

3. Lokacin yankan manne, yi ƙoƙarin yanke kawai manne tare da tube don rage girman yankan manne: kada ku buƙaci manne tare da ɓangarorin maƙasudi kaɗan, in ba haka ba zai shafi ƙimar dawowa.

4. Bayan narke guda biyu ko fiye na manne, yi amfani da bututu komai girman girman kuma canza shi zuwa shafi ɗaya.

5. Maganin da aka kara a cikin sol zai iya zama dan kadan, wanda ya fi dacewa da haɗin DNA zuwa membrane, amma gabaɗaya baya wuce 750ul.

6. Makullin dawo da gel shine don ɗaure DNA zuwa ginshiƙi ta hanyar haɗin gishiri, acidity (cajin) da hydrophobicity na bayani a cikin shafi.Don haka, idan pH na buffer electrophoresis ya yi yawa, 10ul (pH 5.0, 3mol/L NaAC) za a iya ƙara zuwa sol;don mafi kyawun tsangwama kwayoyin DNA akan membrane, ana iya ƙara 30% isopropanol zuwa zafi a cikin ruwa bayan narkar da manne.

7. Kafin ƙara eluent, bar ginshiƙi a zafin jiki na 'yan mintoci kaɗan (kimanin mintuna 10) don cika fitar da ethanol.

8. A ƙarshe, ƙara ƙarancin haske don rage girman farfadowa.Gabaɗaya, ana amfani da 30-50μl eluent don haɓakawa (ba kaɗan ba, in ba haka ba ba zai iya jika membrane ba, wanda bai dace da haɓakawa ba);ɗigon ƙyalli na ƙyalli suna cikin tsakiyar membrane, don cikar DNA ɗin da ke ɗaure ga membrane.

9. Bayan ƙara eluent, ana iya cire shi a cikin wanka na ruwa na digiri 55 na minti 5 ko sanya shi a cikin wanka na ruwa na 50 digiri fiye da minti 10, ko kuma a rufe shi da parafilm a digiri 4 a cikin dare, sa'an nan kuma a sanya shi a tsakiya don farfadowa a rana mai zuwa, sakamakon yana da kyau.

10.Ƙara centrifuged eluate baya zuwa ginshiƙin talla kuma a sake sakewa.

Hanyoyi da ƙayyadaddun hanyoyin don dawo da samfur na PCR

1. Sake amfani da roba na yau da kullun

Idan kuna son dawo da manne, yana da kyau a yi amfani da kit ɗin, wanda ya dace kuma yana da ƙimar dawowa kaɗan kaɗan.Idan da gaske kuna buƙatar dawo da shi da hannu, zaku iya ƙara ƙarar TE sau 3 bayan yanke manne.Bayan narkewa a cikin wanka na ruwa, ana fitar da phenol, phenol/chloroform da tsabta, kuma ethanol yana haɗe.Shi ke nan.

2. DNA dawo da daga low narkewa batu gels

Tsabtace gutsuttsuran DNA Ƙara TE (10 mmol/l Tris-HCl pH8.0, 0.1 mmol/l EDTA) daidai da ƙarar gel, kuma sanya a cikin ruwan wanka na 65 ° C na minti 5 don narkar da gel gaba ɗaya.

Bayan an kawo shi cikin dakin da zafin jiki, daidai da adadin phenol (cikakken da TE, TE an rufe shi a cikin babban Layer, kuma an cire ƙananan Layer na phenol) kuma an haɗa shi a hankali (ba a buƙatar haɗuwa), kuma an sanya shi a 12,000 rpm na minti 3.Maimaita sau 1-2.

Ɗauki mai ƙarfi, ƙara ƙarar 0.1 na 3mol/L sodium acetate (pH 5.2) da ƙarar sau 2.5 na cikakken ethanol don aiwatar da hazo ethanol.Narkar da DNA ɗin da aka tsarkake tare da adadin TE da ya dace, auna abun ciki, kuma shirya don amfani (ana iya amfani da shi don nazarin tsarin tsarin kwayoyin halitta, shirye-shiryen bincike, da dai sauransu).

3. PCR dawowa tare da ƙayyadaddun haɓakawa mai kyau

Idan ƙayyadaddun haɓakawa na PCR yana da kyau, kawai tsarkakewa ne da dawo da samfurin PCR.Kuna iya ƙara 50ug/ml proteinase K zuwa samfurin PCR, digiri 37 na 1h, cirewa sau ɗaya tare da phenol/chloroform, cire sau ɗaya tare da chloroform, kuma ƙara ƙarar 0.1 na supernatant.An dawo da acetate sodium ta hazo tare da 2.5 kundin cikakken ethanol.

Samfura masu alaƙa:

https://www.foreivd.com/pcr-purification-kit-2-product/

https://www.foreivd.com/blood-superdirecttm-pcr-kit-edta-product/