Schistosoma Genome Mapping - call for letters of Support

David Johnston daj at nhm.ac.uk
Mon Jul 17 08:58:56 EST 2000


Dear Collegues,

This email is to update you on important new developments in Schistosoma
genome analysis and to request your support for a major new initiative (see
below). I apologise for the length of this email but this new initiative
needs to be viewed in the context of the other projects.

The Wellcome Trust recently called for proposals for access to the
facilities of the Pathogen Sequencing Unit (PSU) at the Sanger Centre. Of
the 3 areas of support offered, 2 are of direct interest to the Schistosoma
Genome Network and the wider Schisto community

(1) Comparative sequencing projects for pathogens where at least one
representative of a group of pathogens has been sequenced.

(2) Pre-sequencing projects, to provide a basis for sequencing large
pathogen genomes. This could include preparation, gridding and distribution
of accredited large genomic DNA libraries, end-sequencing of large insert
library clones, physical map construction, pilot sequencing of contiguated
genomic regions, EST sequencing for gene discovery and sequence annotation.

On behalf of the Schistosoma Genome Network, I have been in discussion with
the PSU and, as a result of those discussions, we are developing a proposal
that would include the following:

(1) to produce a Happy map of  S. mansoni. Happy mapping is a relatively
new, PCR based way to develop high resolution "genome linkage" maps. It is
analogous to radiation hybrid mapping but uses a panel of DNA samples,
rather than a panel of cell lines and is much quicker and more reliable
than RH mapping. Genomic DNA is fragmented by shearing or irradiation and
separated into pools, each of which contains less than one genome's worth
of DNA. These samples are then screened by PCR  for markers identified by
EST and BAC end sequencing projects. If markers are closely linked, they
are likely to reside on the same fragment and so amplify from the same
sample. Several 10s of thousands of markers can be mapped in parallel this
way. The technique has been used to produce  maps of human chromosome 14,
Dictyostelium chromosome 6 and the complete genome of Cryptosporidium.

(2) to use the Happy map to identify a BAC minimal tile path as a prelude
to genomic sequencing in S. mansoni

(3) to fully sequence 1MB of the S. mansoni genome in order to investigate
genome organisation and verify map construction

 (4) to investigate the use of optical mapping in Schistosoma mansoni

(5) to generate a pilot EST dataset for S. haematobium (for which there is
virtually no sequence data available) and possibly also for another
flatworm (Fasciola, Taenia, Echinococcus or similar) for comparative
purposes. Potentially 15,000 ESTs for each.

(6) Surplus PCR primers from the mapping project would be gridded as
microarrays.

These topics have been selected as a result of recent developments in EST
and genomic sequencing projects (see below) and S. mansoni  - Puerto Rican
strain has been selected as (a) it is one of the most widely distributed
laboratory strains and (b) it is the source of the existing and proposed
BAC libraries.

S. mansoni Genomic Sequencing. The Network has (1) a funded project for
20,000 BAC end seqs (10 MB)and 300KB of fully annotated seq to be
undertaken at CNS, Paris (starting  now and using the Network's existing 8x
coverage BAC library), (2) a project submitted to NIH for US$ 3,000,00 of
earmarked funds for a further 14,000 BAC end seqs (7MB) and 5MB of fully
annotated seq from chromosome 3, plus the construction of new 20x BAC
library (this project to be done at TIGR, we are awaiting news of the
decision) and (3) a preproposal to the NHMRC in Australia for a further
20,000 BAC end seqs (10 MB)and 5MB of fully annotated seq to be undertaken
at the Australian Genome Research Facility.

EST sequencing. In addition to the existing data sets of 16,000 S. mansoni
and  2,000 S. japonicum ESTs (1) Sao Paulo and Minas Gerias States in
Brazil are proposing to commit US$ 1,500,00 each to establish a networked
genome centre in Minas Gerias, like the ONCA project in Sao Paulo (which
was responsible for the 2.7Mb- Xylella fastidiosa genome that was published
in Nature last week) and to use this to generate, amonst other things,
large numbers of S. mansoni ESTs  (2)  The Chinese National Human Genome
Center at Shanghai and the Institute of Genetics, Chinese Academy of
Science (IG/CAS) in Beijing have just announced a collaboration with the
Institute of Parasitic Diseases in Shanghai to generate 100,000 ESTs from
adult male and female S. japonicum by the end of 2000.

Large scale  EST and genome sequencing projects are, therefore, underway.
Large scale mapping is the missing piece of the jigsaw and the PSU is one
of the few Centres to offer access to such a  service. We need a physical
map  because the Schistosoma genome is so large, contains large amounts of
repetitive DNA and cannot be partitioned by PFGE or cytometry. Therefore, a
shotgun approach to obtain full genome sequence would be unlikely to
succeed (and also would be difficult to justify at this time). Future
efforts to produce contiguous sequence for selected regions of the genome
will, therefore, depend on having a physical map to guide clone selection
and assembly.

At the same time, the new EST initiatives are likely to identify virtually
all of the genes expressed in adult S. mansoni and S. japonicum. Thus,  if
a gene identified in a new species (like haematobium) has a homolog in
mansoni or japonicum, that homolog is very likely to be already represented
in the datasets, allowing extensive comparative analysis. For example,
comparison of  divergence rates  will identify rapidly evolving genes.
These are likely to be the genes under greatest selection pressure, which
may represent potential weaknesses in the parasite to which drugs or
vaccines could be targeted . In conjunction with the S. mansoni physical
map, the haematobium ESTs should also permit "cloning-by-synteny"
approaches to be developed; if housekeeping genes flank an interesting gene
in one species, they may well do so in another, allowing it to be isolated
by long range PCR.

******************************************

HOW YOU CAN HELP

In order for this application to be successful, we must demonstrate that
there is appropriate UK and international research community support for
the project.

I am therefore writing to request that you send to me, letters or faxes in
support  of the project and of the species / strain selected. If at all
possible, please reply by fax, on headed notepaper, rather than by email .
The preproposal must be submitted by mid August, so an early response would
be appreciated.

A suggested format for the letter is below, please feel free to use this
format "as is", or to edit it, or to provide a letter of your own format.


Thank you, in advance, for your support.

David A Johnston,

on behalf of the WHO Schistosoma Genome Network.

*******************************************
suggested format for the letter

Dr David Johnston,
Secretary, WHO Schistosoma Genome Network,
Biomedical Parasitology Division,
The Natural History Museum,
London.

Dear Dr Johnston,

I am writing to express my support for your application to the Wellcome
Trust's Beowulf scheme for a project to develop a physical map of the
Schistosoma mansoni genome by an integrated "HAPPY' mapping and BAC tile
path approach. The completed map, together with data revealed by the
proposed 1MB of contiguous  genomic sequence will be a valuable resource
for my research work, and for the community as a whole, providing an
essential framework for future genome sequencing efforts, contributing to
our understanding of genome organisation and facilitating the isolation and
analysis of regions of  the genome of special interest (including the sex
chromosomes). At the same time, a pilot EST project for S. haematobium will
provide essential data for this poorly characterised, but important, human
pathogen and will allow comparative analysis, including the identification
of rapidly evolving genes. In conjunction with the S. mansoni physical map,
it should also permit "cloning-by-synteny" approaches to be developed for
this and other species.

I confirm that S. mansoni, Puerto Rican strain is the logical choice of
organism for the maping project as it is one of the most widely distributed
laboratory strains (******please say if you actually keep it in your own
lab*******) and as it is the source of the existing and proposed new BAC
libraries that will generate the genome survey sequences to be used as
markers on the map.

Yours sincerely,

*******************************************

David A. Johnston,
Secretary to the WHO Schistosoma Genome Network,
Biomedical Parasitology Division,
Dept. of Zoology,
The Natural History Museum,
Cromwell Road, London SW7 5BD, England, UK.

***PLEASE NOTE NEW PHONE / FAX NUMBERS***

Tel: 020-7942-5566/5008/5152
Fax: 020-7942-5347
(from outside the UK: 44-20-7942 ****)

***PLEASE NOTE NEW PHONE / FAX NUMBERS***

eMail daj at nhm.ac.uk

http://www.nhm.ac.uk/hosted_sites/schisto/

The  Biomedical Parasitology Division is a WHO Collaborating Centre for the
identification of schistosomes and their snail hosts.








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