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Hello,
I am trying to use trueseq DNA PCR free for library prep. I used diagenode bipruptor and not covaris (as mentioned in the protocol). I used settings for 350 but more than half of my fragments are below 200bp. I used 100ul (20ng/ul) for shearing.
Can I combine, say 150ul of sheared DNA, pcr purify it, and use all as input. (Basically I will be using around 2.5ug as input (the protocol suggestion is 1ug). Will 2.5 be too much for end repair step to cope, and repair all of them?
Does anyone has any experience with this?
I tried doing the size selection step (followed the protocol) before the end repair but looks like I lost a lot of DNA during this.
Any suggestions?
I am trying to use trueseq DNA PCR free for library prep. I used diagenode bipruptor and not covaris (as mentioned in the protocol). I used settings for 350 but more than half of my fragments are below 200bp. I used 100ul (20ng/ul) for shearing.
Can I combine, say 150ul of sheared DNA, pcr purify it, and use all as input. (Basically I will be using around 2.5ug as input (the protocol suggestion is 1ug). Will 2.5 be too much for end repair step to cope, and repair all of them?
Does anyone has any experience with this?
I tried doing the size selection step (followed the protocol) before the end repair but looks like I lost a lot of DNA during this.
Any suggestions?