After a MSA using clustal, can I load the alignment in FASTA format to FastML to get a reliable ancestor protein sequence prediction?
Yes, you can use FastML to get reliable ancestor sequences. I should note that the previous steps leading to this need to be performed very carefully, i.e., your alignment and tree need to be constructed correctly. The REAP protocol is pretty good at discussing the steps involved (paywall).
FastML itself can be annoying to use. Occasionally the server will not work (for whatever reason) and at other times very minor formatting issues in your supplied alignment or tree will cause it to break (and the server won't always tell you what the issue is).
An alternative is CODEML in the PAML package... although this is incredibly difficult to use and the documentation is crap hard to interpret.
I can expand on the sequence of steps that I believe need to be taken for ASR (formatting tricks, programs to use, etc.) if required.
N1 is the oldest ancestor. After that, all other internal nodes of the phylogenetic tree are labelled in order ....N2, N3...etc. N2 and N3 for example would be children of N1. Extant sequences should maintain their original labels. I believe FastML provides a newick tree in one of the output files with the ancestor labeling included...so plotting that should give you a more `visual' idea of how the nodes are labeled.
Hi
I have a few more doubts. Which kind of output should we consider regarding FASTML? Is it joint reconstruction file or marginal construction with/without indels?
One more thing - I have used an outgroup while making the tree. Will this interfere with the accurate ASR?
Please answer...
Hmm good question. Depends on what your ultimate goal is, but typically you will just use the joint reconstruction (If you read up on the theory on the difference between joint and marginal this will make more sense). The marginal reconstruction is useful if you want to compare against the joint and get a sense of what positions were difficult for Fastml to infer (at least I think so?)....e.g. a position in an ancestor could be 50% likely to be a 'G' and 50% likely a 'C', rather than 90% likely.
And yes, you are meant to use an outgroup (depending on what you are reconstructing). I forgot to mention that before.
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version 2.3.6
Hello,
I have a question: how to calculate the difference in log-likelihood between the most likely ancestral sequence at node N1 and the 100th most likely sequence??
thanks in advance
laura