xMAN commandline

Here is the xMAN how to from Tao Liu


Text formatted bpmap may not be processed by MAT directly. You need to convert your text bpmap to a so-called 'plainseq' format. Please type 'xMAN' without any argument to see the usage for detail.

And here is the commandline used to generate bpmap for fruitfly:

xMAN -f bpmap -i Dm_tiling2_MR_v01.bpmap -s Dm_tiling2_MR_v01.stderr -o Dm_tiling2_MR_v01_dm3 --nro=Dm_tiling2_MR_v01_dm3_all.NR --maxSeqCopy=10 --ProbeResolution=35 dm3/all/*.fa

The input file is a binary bpmap downloaded from affy website. And I put all the genome sequence files in dm3/all/ directory. Please remember to add '-nro' argument. After the run, the file 'Dm_tiling2_MR_v01_dm3_all.NR.bpmap' can be used by MAT.

Making MAT RepeatLib

Download the repeat_mask.txt, simple_repeat.txt and segment_dups.txt from UCSC

and run

python Rep.py -m repeat_mask.txt simple_repeat.txt segment_dups.txt

The Rep.py is available at the MAT lib subdirectory of the MAT install.

For the genomes like Drosophila melanogaster, for which UCSC doesn't provide Segment Duplication file. The repeat library can be generated by

1) remove all the instances of usage of "segdup" from Rep.py file
2) remove the original Rep.py and Rep.pyc files form MAT compile.
3) Recompile MAT.

RepeatLib and the modified Rep.py files are also available on request from me.
parantu dot shah at gmail dot com

Installing python module locally

python setup.py install --prefix=/my/fav/dir

BLAT PSL coordinates

.psl files

A .psl file describes a series of alignments in a dense easily parsed text format. It begins with a five line header which describes each field. Following this is one line for each alignment with a tab between each field. The fields are describe below in a format suitable for many relational databases.

matches int unsigned , # Number of bases that match that aren't repeats

misMatches int unsigned , # Number of bases that don't match

repMatches int unsigned , # Number of bases that match but are part of repeats

nCount int unsigned , # Number of 'N' bases

qNumInsert int unsigned , # Number of inserts in query

qBaseInsert int unsigned , # Number of bases inserted in query

tNumInsert int unsigned , # Number of inserts in target

tBaseInsert int unsigned , # Number of bases inserted in target

strand char(2) , # + or - for query strand, optionally followed by + or – for target strand

qName varchar(255) , # Query sequence name

qSize int unsigned , # Query sequence size

qStart int unsigned , # Alignment start position in query

qEnd int unsigned , # Alignment end position in query

tName varchar(255) , # Target sequence name

tSize int unsigned , # Target sequence size

tStart int unsigned , # Alignment start position in target

tEnd int unsigned , # Alignment end position in target

blockCount int unsigned , # Number of blocks in alignment. A block contains no gaps.

blockSizes longblob , # Size of each block in a comma separated list

qStarts longblob , # Start of each block in query in a comma separated list

tStarts longblob , # Start of each block in target in a comma separated list

In general the coordinates in psl files are “zero based half open.” The first base in a sequence is numbered zero rather than one. When representing a range the end coordinate is not included in the range. Thus the first 100 bases of a sequence are represented as 0-100, and the second 100 bases are represented as 100-200. There is a another little unusual feature in the .psl format. It has to do with how coordinates are handled on the negative strand. In the qStart/qEnd fields the coordinates are where it matches from the point of view of the forward strand (even when the match is on the reverse strand). However on the qStarts[] list, the coordinates are reversed.



Here's an example of a 30-mer that has 2 blocks that align on the minus strand and 2 blocks on the plus strand (this sort of stuff happens in real life in response to assembly errors sometimes).

0 1 2 3 tens position in query
0123456789012345678901234567890 ones position in query
++++ +++++ plus strand alignment on query
-------- ---------- minus strand alignment on query

Plus strand:
qStart 12 qEnd 31 blockSizes 4,5 qStarts 12,26
Minus strand:
qStart 4 qEnd 26 blockSizes 10,8 qStarts 5,19

Essentially the minus strand blockSizes and qStarts are what you would get if you reverse complemented the query.However the qStart and qEnd are non-reversed. To get from one to the other:
qStart = qSize - revQEnd
qEnd = qSize - revQStart

Running BLAT with maximum sensitivity

nohup /home/shah/local_progs/blat_dir/blat -fine -repMatch=1000000 -minIdentity=100 /home/shah/drosophila_annotations/dmel3_apr_2006/dmel3_apr_2006.fasta agilent_dm2_probe_sequences_5_215453.fasta agilent_dm2_probe_sequences_5_215453.psl &

explanation :

-fine
-repMatch=1000000 - number when tiles are considered overused
-minIdentity=100 - pick identical matches

e.g. for mapping probes to the reference genomes.

Installing R package tar.gz

use

R CMD INSTALL pkg.tar.gz


or within R session

install.packages("pkg.tar.gz", type="source", repos=NULL)

Running xMAN for Affy probe mapping

According to Drosophila Tiling 2.0R array design and library file description Affy tiling2.0R BPMAP file may contain probes that map to genomes multiple times, which may be derived from repeated gene families and other sequences that were not repeat masked.

Existence of probes that map to genomes multiple times lead to inflated p-values during the subsequent analysis and lead to false-positives. Also, remapping the probes to a newer genome assembly is desirable sometimes

Thus, it is important to remap the BPMAP to the genomes.
xMAN is available from Shirley Liu's lab.

Command line while running xMAN may look like

First try:

xMAN -f bpmap -i Dm_tiling2_MR_v01.bpmap -o Dm_tiling2_MR_v01_xman_nr.bpmap --nro=Dm_tiling2_MR_v01_xman_nro.bpmap --maxSeqCopy=10 chr2L.fa chr3L.fa chr2RHet.fa chr3RHet.fa chrUextra.fa chr4.fa chrX.fa chr2LHet.fa chr3LHet.fa chrM.fa chrXHet.fa chr2R.fa chr3R.fa chrU.fa chrYHet.fa

Generating a Affymatrix "tpmap" file for TiMAT

R Code that uses the Bioconductor module Affxparser.

Note: The code has a bug and in normal circumstances returns an error

> writeTpmap(filename="Dm_tiling2_MR_v01.tpmap", bpmaplist=bpmapr)
> Error in as.vector(y) : argument "y" is missing, with no default

The "y" can be fixed as follows

> y <- readBpmap("Dm_tiling2_MR_v01.bpmap")
> writeTpmap(filename="Dm_tiling2_MR_v01.tpmap", bpmaplist='y', verbose = as.integer(0))

This seems to work but doesn't print anything into the tpmap file due to some "sequence skipping".

I will try the Affymatrix windows based program next.

me vs. reggaeton

Doing a PhD in Germany was a wonderful experience. Those 4.5 yrs taught me German and (mostly reading the literature) improved my English .. My proficiency over Latin based languages didn't improve !! Zero in French and in Spanish .. despite boasting a PhD from Autonoma De Madrid !!!

Now US is making sure that I learn some Spanish ... nope no spanish girl friends!! My wife will kill me if I do it. Its mostly the reggaeton ... from Don Omar, Daddy Yankee, Wishin Y Yandel , and others. I like the beats even though they are far away from Rammstein :D ... If you ask "Who is Don Omar" , then give a You tube search !!! Highly recommanded.

Protein 3D structure analysis

I was trying to find today programs for analyzing protein 3D structures .. A rare art
new generation of Bioinformaticians may never learnt. Especially, I wanted to calculate secondary structure, solvent accessibility, hydrogen bonds, torsion angles and residue packings from protein 3D structures.

Any ways, the old programs viz. Comparer, JOY suite (people believe its no fun at all running it) at the University of Cambridge and WHAT IF are still under copyrights that you still have to FAX them to get a license. (aka long wait .....)

I finally downloaded DSSP (CMBI version), STRIDE (from EMBL/EBI) and PSA (sali lab). Also, I will try VADAR suite of algorithms from Canada and PROMOTIF (I never used it before though) to get most of the required information.

Let's see where these guys take me !! Anybody who want to write Bioperl modules for them :D !!!

Talibanization, Pakistan and Global Consequences

The government of President Mussharraf is under unprecidented crisis. No words minced. As the preceding dictators of Pakistan never faced a crisis akin to what Pres. Mush is facing. Pres. Bhutto and Zia-Ul- Haq survived and ruled by encouraging increasing islamization of Pakistan society. It was a requirement to fight the communist russians in Aghanistans and that bred the Mujahhidins .. and Talibans. With the end of the cold war and collaspe of Soviet union, the ground reality suddenly changed. And with September 11, 2001 the Pakistan government is now required to fight against the militias they helped training with US support.

With US/NATO troups in Afghanistan, the Talibans filtered into frontier regions in Pakistan. Pres. Mush. made a pact with the devil and allowed the law of sharia in those provinces. But now the Lala Masjid (Red Mosque) episode in Islamabad suggests that he is just a helpless president, unless he plays a master political stroke. Else, under fire from both sides, the inteligencia with Justice Chowdhry and militia with Red mosque cleric brothers, he might be loosing the power soon. Let's see how far Allah, Army and America can taqke him.

General in the line of fire with Stewart

It's now a relatively old one ... but a must see for all people who wants to learn the art of tight rop walking. Three salutes to the General.



Or you can see this on
http://www.youtube.com/watch?v=GJWwmNCMVpo

Does Japanese Bioinformatics servers work?

This is a general frustration for me and a couple of friends of mine whether japanese Bioinformatics server ever work for outsiders? I am not puting the famous servers into these categories but a few examples are

http://medaka.dsp.jst.go.jp/MEPD
http://timpani.genome.ad.jp

Does any body join my frustration?

My 2 cents on Israel- Hizbollah 2006

1) Hizbollah started it by kidnapping two soldiers. In past such kidnapping has been used for bargaining their prisoners out by both Hamas (Palestin) and Hezbollah.

However, the motive of Hizbollah this time is different after the voluntary evacuations of jewish population from Gaza strip and new prime minister at helm in Isreal.

2) Obviously, the heavy handed approach of prime minister Olmert is being critisized as too much or killing of innocent citizens.

But it is important to disarm hizbollah for various reasons.

3)
- Hizbollah is heavily armed !! militant group which can cross national borders (without the government backing) and plunge the country in to a war.
- it doesn't represent a soverign government and hence has no rights to be armed !!!.
- It is influenced by Syria and Iran which has its own agendas in supporting it.
- It has clearly islamic agenda which can endanger the christian minority and democracy in Lebanon and it has stated goal of destroying Israel.
- Moreover, they are fighting while hiding among civilians.

4) Diplomacy - France is going back its former protectorate just like English sitting in Basra and Afghanistan.

Reservation and affirmative action - some thoughts on implimentation

The idea is to discuss some feasible ways to impliment affirmative
action and also let the talents flurish. Moreover, the ways to tackle
"creamy layer" inside the SC/ST/OBC community who are taking unfair
advantage of the system is considered.

1) Affirmative action is necessary.

The socially and economically backward class has a right to join the
forefront of the society and taste the fruits of progress country has
made in last 50 yrs. This is important for a balanced growth and
healthy society.

2) Affirmative action should be implemented on the basis of economic
well-being and edicational level

Indian society has been troubled enough by the caste system. As a
secular nation implementing any steps on the basis of caste or
religion is against our constitution, so is the quotas based on caste.
It will divide the nation.

Instead, it is the level of education and economic well-being that
should be taken into consideration.

Students whose parents are not educated up to a college graduate level
and not earning enough should be give "free" school and college
education regardless of their caste or religion.
On the other hand, How can a person who finished an MBBS or a BE
(technology) could be considered backward class? Thus, quotas in
higher education (post graduate studies) should be restricted (see
below).

3) Quotas on job:

It is possible that even in the 21st century the educated person is
discriminated against because of their caste, but it is getting less
and less likely due to healthy private sector and cut-throat
compitition where talent is the only way. If the government wishes so
- they can have quotas in the job in govt. sector. Decision making in
Private enterprises should be private and should not be pressured by
Govt.

4) Restricting the creamy layer.

It is impossible to have everybody taking advantage of the affirmative
action at the same time and it is also impossible to stop some
families from falling to poverty. So it is very hard to put a stop as
to when affirmative action should end. But it is equally important to
stop the creamy layer from taking unfair advantage of the quota
system.

Instead, an SC/ST/OBC candidate should be given choices. Once in his
life time he can choose between having the affirmative action in
favour of him for Post graduate studies or his first job. The
affirmative action should stop there.
Similaraly children of educated parents (college graduate) should not
be able to take advantage of affirmative action after 3 generations of
their forefathers having utilized it . Though, economic aid should be
allowed to all those in need.

5) Caste and religion based advantages should be strictly forbidden
and constitution should be restored to remove reservation.

By the rights given to us in our constitution all indian are equal
irrespective of their caste or religion and this should be respected.
Reservation was a temporary measure and constitution was amended for
allowing it. Thus, it should be restored. Again, affirmative action is
important but it should not be the birth right on any individual.

The logic of mobocracy: Mandal II

This is to sound my protest against the newly introduced political move by Congress led UPA government in India.

First, let us review some points that are time and again cited in favour of reservation.

1) The "dalits" (SC/ST/BC and now OBC) have been kept at the periphery of the society and never given the polictical and social rights that they deserve
2) Their financial and educational levels are low in comparision to the "upper" classes, especially in the rural area.
3) The government need to improve their status by affirmative action - i.e. reserving upto 50% of seats in the education as well as job sector.

Above three points makes the assumption that social and financial well-being is tied with "caste" i.e. it is desided to which household you were born.

I should start with reminding the "Right to equality" that is given to all the citizens of India by our constitutions. I am aware that our constitution had been amended to introduce the reservation. But I still consier the reservation as UNCONSTITUTIONAL and reservation as an illigal step against the right to equality.

Second, I am not against the affirmative action. But instead of tying it with a particular caste (i.e. birth right) it should be given to the all economically and socially backward members of the society and it should be stopped after 1 or 2 generation of house hold has utilized their opportunities.

We all know that SC/ST enjoyes the reservation since last 50 years or so in every aspect of social life. So if reservation is working then they should have been the prosperous members of the society and after 3 or more generations that they have enjoyed this right it is a right time to remove reservation as given to them. However, if they still reqiure the support of reservation in order to prosper, that means that reservation given to particular caste has failed and the measure should not be applied to the larger section of the society as it will fail while hurting growth of rightful candidates.

Though, we all know that it such reservation has resulted in a creamy layer across SC/ST classes in terms of political and job markets. It hasnot changed the situation at the grass root level.

Why it is a political master punch from Congress?

Let us remember some history. Not so long ago Mr. VP Singh, then the prime minister of india desided to introduce the steps as Mandal panch required. This resulted in wide spread protests and self-immolation. It finally resulted in fall of the government. The next government Congress supported for a while and then pulled back its support, which lead to the political come back for the party and the government of Mr. PV Narsimha Rao. The reason being strong support for congress at the grass root level. The situation hasn't changed much. The Congress came back to power after Mr. Vajpayee's BJP govt. was defeated by support of the indian villages. So now, by introducing this contraversial moves (which no political parties can oppose !!) congress is making sure for a next strong government and a very strong vote bank. Thus, changing democracy to mobocracy.

Now discussion about an unrelated topic that gives the same message. The controversial movie (and book) "Da Vinci Code" was banned in
christian majority state of Nagaland citing its potential for political disturbance but the same reason was given for banning it in Punjab, where christians are in minority. It not only shows political hand on the neck of freedom of expression but again the "mobocracy" at work.

We, as Indians has to be aware of this poison else the dream of being 21st century suporpower would be drowned in the Indian ocean.

Got the Adsence running

Finally got the AdSence running today ... that was easy ... looks like I am finally getting hand of computers ....
Don't get impressed ... just go to Google As sense and copy paste the code at proper places.
Now let's see how fast the $$$ turn up !!!

Identifying good targets for making antibodies.

The idea is to identify regions that are highly antigenic and and specific to the protein under study. Looking at a multi species view that would mean that regions that are highly antigenic (immunogenic), conserved across orthologues but not conseved amond paralogs (homologues proteins in the same species). Also, the problem of cross reactivity could occur by identical stretches in non-related species. Thus, a blast is necessary on the entire genome to rule out the similar regions.

Finding antigenic regions
----------------------

Now in order that antibodies can react specifically to the regions in proteins, follwing properties are necessary.

1) The region should be solvent accessible
2) The region should be hydrophilic
3) The region should have a specific secondary/tertiary structure.

Such regions can be divided into two - 1) continuous stretches along the protein sequence and 2) Non-continuous stretches along the protein sequence but continuous in the tertiary structure (protein surface)

Other desired structural properties are - 1) presence of a proline (possibility of a cis confirmation), beta turn, and so forth.
3D structures would help identifying non-contiguous stretches.

Methods:
--------

1) Blast again Genome as well as PDB
2) SMART/Pfam
3) hydrophobicity/hydrophilicity index and
4) Secondary structure and slovent acessibility prediction (PHD)
5) Turn prediction and soforth.


Any suggestions are well come

The Big idea !!

The idea is to start a discussio blog about high-throughput molecular biology .. sharing some tips and trics of the trade.
I work on Bioinformatics so my posts will mostly be leaning towards computational biology.

Ofcourse other discussions are allowed !!
Check for more in future !!!