求一个好用的给genotyping PCR 的DNA isolation protocol# Biology - 生物学
b*n
1 楼
目前用的是Jackson lab的quick dirty protocol, 如下：
NaOH extraction (quick "dirty" DNA preparation). Reference: Truett GE et al.
2000. Biotechniques 29(1):52-54
Cut 2mm of tail and place into an Eppendorf tube or 96-well plate
Add 75ul 25mM NaOH / 0.2 mM EDTA
Place in thermocycler at 98ºC for 1 hour, then reduce the
temperature to 15°C until ready to proceed to the next step
Add 75ul of 40 mM Tris HCl (pH 5.5)
Centrifuge at 4000rpm for 3 minutes
Take an aliquot for PCR (use 2 ul undiluted, or 2 ul of a 1:100 dilution
/reaction)
感觉不是很爽，有一些tough的genotyping作不出来。
求推荐一个好用的替代，感谢！
NaOH extraction (quick "dirty" DNA preparation). Reference: Truett GE et al.
2000. Biotechniques 29(1):52-54
Cut 2mm of tail and place into an Eppendorf tube or 96-well plate
Add 75ul 25mM NaOH / 0.2 mM EDTA
Place in thermocycler at 98ºC for 1 hour, then reduce the
temperature to 15°C until ready to proceed to the next step
Add 75ul of 40 mM Tris HCl (pH 5.5)
Centrifuge at 4000rpm for 3 minutes
Take an aliquot for PCR (use 2 ul undiluted, or 2 ul of a 1:100 dilution
/reaction)
感觉不是很爽，有一些tough的genotyping作不出来。
求推荐一个好用的替代，感谢！