b*******n
发帖数: 605 | 1 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！ | D*a
发帖数: 6830 | | n***w
发帖数: 2405 | 3 DirectPCR lysis reagent (Viagen Biotech 102-T) and proteinase K (Viagen
Biotech 501-PK).
100ul lysis reagent + 2.5ul proteinase K, 55 degree O/N (>2hrs or more), 95
degree inactivate 15min
ready to go.
Sometimes you may need to consider DNA purification. | c*********t
发帖数: 340 | |
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