2 × PCR Premix (ifite irangi) HYA442
2 × PCR Master Mix irimo Taq DNA Polymerase, dNTPs, n'ibindi bice bikenerwa na PCR. Master Mix ihoraho amezi 3 kuri 4℃ hamwe n'ibikoresho byacu bihamye byo gupima. Umuti wa pre-mix wakozwe neza kuri PCR isanzwe kandi witeguye gukoreshwa wongeyeho ADN template na primers. Ibikoresho bya PCR bishobora gushyirwamo electrophoresis hakoreshejwe irangi ry'ubururu rya bromophenol ryashyizwemo. Ibikoresho bya amplified birimo 3 '-dA protrusion kandi bishobora guhindurwa byoroshye muri T vector. 2 × PCR Master Mix yoroshya uburyo bwa PCR kandi ikagabanya ubwandu.
Ibice bigize
| Ibice bigize | Ingano-1 | Ingano-2 | Ingano-3 |
| 2 × PCR Premix (ifite irangi) | 5ml | mL 1 × 15 | mL 1 × 50 |
Ububiko
Bika ahantu hadakoreshejwe -20°C mu gihe cy'imyaka 2.
Amabwiriza
1. Sisitemu yo Gusubiza
| Ibice bigize | Ingano |
| 2 × PCR Premix (ifite irangi) | 25 uL |
| Igikoresho cyo gushushanya imbere (10 μM) | 2 µL |
| Igikoresho cyo gushushanya inyuma (10 μM) | 2 µL |
| Icyitegererezo cya ADN* | Ihinduka |
| ddH2IKI | Kugeza kuri 50 µL |
* Ikoreshwa ry'icyitegererezo: 50-200 ng ADN genomic; 0.1-10 ng ADN ya plasmid
2. Gahunda yo Gusubiza Ibibazo
| Intambwe | Ubushyuhe | Igihe | Ingendo |
| Guhinduka kw'ibanze | 94 ℃ | Iminota 5 | 1 |
| Guhinduka kw'ingufu | 94 ℃ | Amasegonda 30 | 35 |
| Annealing | 50-60 ℃ | Amasegonda 30 | |
| Kwagura | 72 ℃ | Amasegonda 30-60/kb | |
| Kwagura kwa nyuma | 72 ℃ | Iminota 10 | 1 |
| Gufata | 4 ℃ | - | 1 |
1. Mg2+ ikigero: Iki gicuruzwa kirimo 3 mM ya MgCl2, ikwiriye cyane mu mikorere myinshi ya PCR.
2. Ubushyuhe bwo gufunga: Reba agaciro ka Tm k’ibishushanyo mbonera by’ibishushanyo mbonera. Ubushyuhe bwo gufunga bushobora gushyirwa kuri 2-5°C munsi y’agaciro ka primer.
3. Igihe cyo kongera igihe: Kugira ngo hamenyekane molekile, ni ngombwa ko habaho amasegonda 30/kb. Kugira ngo hakorwe gene, ni ngombwa ko habaho amasegonda 60/kb.
Inyandiko
1. Ibikoresho bya PCR bifite 2 × PCR Master Mix ntibikwiriye polyacrylamide gel electrophoresis.
2. Ku bw'umutekano wawe n'ubuzima bwawe, mbara amakoti yo muri laboratwari n'uturindantoki dukoreshwa rimwe mu gihe cyo kubaga.
3. Ikoreshwa mu bushakashatsi gusa!