very bad signals after sequencing
Posted 07 June 2010 - 11:23 PM
plasmid: 2 ul (>200 ng)
bigdye (mixed with buffer): 2 ul
primer (1 mM): 4 ul
H2O: up to 10 ul
and PCR is
96C 1 min;
96C 10s,50C 5s,60C 4 min for 40 cycles.
and then I use ethanol/NaAc precipatation and resolve the DNA with formadine. and this protocol worked well before and I dont know why this time it didnot work.
Posted 07 June 2010 - 11:25 PM
Posted 12 June 2010 - 12:26 PM
Posted 18 June 2010 - 08:41 AM
if it was weak then you have a lot of noise.
if it was strong then you may be looking at multiple sequences.
if multiple sequence then you either have more than one primer, more than one priming site or more than one species of plasmid.
talent does what it can
genius does what it must
i used to do what i got paid to do
Posted 30 June 2010 - 07:50 PM
I put the same sample on another sequencer, and this signal pattern never appear again. although I still didnt get 100% clear result, the data is better than that from the previous sequencer. seems like the sequencer didnt work well.
Posted 04 August 2010 - 04:03 AM