Download Exercises

1. Neighbor-joining
The following file contains sequences from precomputed psi-blast search for the human estrogen
receptor alpha (ESR1_HUMAN) against nrdb30 database from the PairsDB database.
Hormone_receptors.fasta
Align the sequences in hormone_receptros.fasta using muscle, save the file in fasta format.
Convert the file from fasta to meg format using Mega.
File > Convert to MEGA format ...
In order for Mega to parse your alignment correctly. You will need to remove comma characters
(,) from the sequence names using Mega text editor. Use Mega text editor to replace, with a blank
space. Make sure all comma characters are replaced by selecting:
Replace > Scope > Global
Phylogenetic analysis
Compute a Neighbour-Joining tree for your alignment. Use the JTT substitution matrix in
distance computation and gamma correction with gamma parameter value 2.4 (Molecular
Evolution and Phylogenetics, Nei & Kumar 2000).
Phylogeny > Construct phylogeny > Neighbour-Joining
Model > Amino acid > JTT matrix
Bootstrap test
Compute bootstrap test using 100 replicates.
Typically it is recommended to perform at least 500 replicates but in order to make computation
faster will use only 100 replicates here.
Identify and name the following subfamilies in the tree. Use:
Right mouse button > selected subtree
Estrogen receptor αs
Estrogen receptor βs
Androgen receptors
Progesterone receptors
Estrogen like pseudogenes (ERRs)
Change the colour of the subfamily branches from:
Subtree > Draw options
Compress the retinoid receptor family branch and define it as the outgroup.
Identify and low confidence nodes that have bootsrap values below 70 %
Q 1. Copy paste the tree image below.
2. Phylogenomic functional annotation
Using the hormone receptor subfamily sequences from the previous exercise as reference, predict
the function of these putative hormone receptor sequences (seq1, seq2) predicted from the
hedgehog genome.
Add sequences seq1 and seq2 to the sequence file and repeat the analysis procedure in exercise 1.
Find out which subfamily do these cluster in. Remove retinoid family sequences from the analysis.
Hints:
Due to incomplete genome assembly parts of the protein sequence are uncertain and
marked with X. When opening your meg file specify the symbol for missing data to
be capital X.
>Seq1
PSSESRRQAGRERLASSGDKGSLSMESAKETRYCAVCNDYASGYHYGVWSCEGCKAFFKR
SIQGHNDYMCPATNQCTIDKNRRKSCQACRLRKCYEVGMMKGGIRKDRRGGRMLKHKRQR
DDGEARSESGPSGDMRATTLWPSPLVIKHTKKNSPALSLTAEQMVSALLEAEPPFIYSEY
DPTRPFSEASMMGLLTNLADRELVHMINWAKRVPXXXXXXXXXXXXXXXXXXXXXXXXXX
XXXXXXXXXXXXXXXNLLLDXXXXXXXXXXXXXXXXXXXXXXXXXXXXXXXXXXXXXXXX
XXXXXXVYTFLSSTLKSLEEKDHIHRVLDKITDTLIHLMAKAGLTLQQQHRRLAQLLLIL
SHIRHMSNKGMEHLYSMKCKNVVPLYDLLLEMLDAHRLHAPANRGAAATMEEMSQGPLAT
NGQASHSLQTYYITGEAENFPTI
>Seq2
EALKRKVSRSSCASPVTSPSTKKDAHFCAVCSDYASGYHYGVWSCEGCKAFFKRSIQXXX
XXXXXXXXXXXIDKNRRKSCQACRLRKCYEVGMVKCGSRRERCGYRILRRQRNSDEQLHC
LSKAKKNGGHVTRVKELLLNSLSPEQLVLTLLEAEPPNVLVSRPNTPFTEASMMMSLTKL
ADKELVHMIGWAKKIPXXXXXXXXXXXXXXXXXXXXXXXXXXXXXXXXXXXXXIFAPDLV
LDRDEGKCVEGILEIFDMLLATTSRFRELKLQHKEYLCVKAMILLNSSMYPLAAATQEGE
SSQKLTHLLNAVTDALVWVIAKSGISSQQQSVRLANLLMLLSHVRHASNKGMEHLLSMKC
KNVVPVYDLLLEMLNAHTLRGYKSSVTRSECSPAEDSKSKEGSHNPQSQ
Q 2.1 Copy paste the tree image below.
Q 2.2 Identify the subfamily and function for seq1 and seq2