2006-07-13 21:43:28 · 6 answers · asked by Anonymous in Science & Mathematics ➔ Biology
I have managed to PCR up to 4.4kb without too much difficulty. I used 68C extension temp, and for a product that was a bit difficult to isolate and kept getting smaller products instead, using 2.5% or 5% DMSO in the reaction seemed to help. I haven't tried bigger products than that. And I used bog standard Taq from Bioline, but added 4ul of Pfu to the new 50ul tube of Taq.
2006-07-14 09:12:06 · answer #1 · answered by Rotifer 5 · 1⤊ 0⤋
be very careful at handling them cause if you really have to use that length, they break easily. Due to the size it is not worth doing it because dna pairs at a 45-50% homolgy, so you will get pairing of almost anything, a trick would be increase the salt concentration in the buffer liquids or lower the taq poly and see if it works the way you want it.
2006-07-13 22:06:06 · answer #2 · answered by Prof. Hubert Farnsworth 4 · 0⤊ 0⤋
There are commercially available TAQ that are designed and optimized for longer amplicons. I think Stratagene has some.
2006-07-14 05:40:02 · answer #3 · answered by pliu428 2 · 0⤊ 0⤋
Boil for 20 minutes with a stock cube and tomatoes, and serve with rice.
2006-07-13 21:48:32 · answer #4 · answered by arnold 3 · 0⤊ 0⤋
It depends on your template. High or low complexity? bacterial? human genomic? GC rich?
Check the following link for some protocols and tips.
2006-07-13 22:05:28 · answer #5 · answered by Slackenerny 4 · 0⤊ 0⤋
grilled
2006-07-14 08:40:39 · answer #6 · answered by GRUMPY /UK 5 · 0⤊ 0⤋