course in YAC analysis
Geordie +1865 740 011
ayoung at vax.oxford.ac.uk
Mon Mar 6 07:21:36 EST 1995
WELLCOME SUMMER SCHOOLS : FIFTHTEENTH ADVANCED COURSE
HUMAN GENOME ANALYSIS : FROM YAC TO GENE
27th July - 4th August 1995
Division of Biochemistry & Molecular Biology, United Medical and Dental
Schools of Guy's and St Thomas's Hospitals, Guy's Campus, London
All email enquiries to: wss at umds.ac.uk
An intensive laboratory course to introduce participants to YAC and cosmid
contig assembly, clone characterisation and gene hunting.
1. Analysis of YAC clones
A range of methods for analysis of individual YAC clones will be covered.
These include the preparation of YAC plugs, gel purification of artificial
chromosomes, experiments to recover sequences from within and at the ends of
YAC clones for further studies.
2. Subcloning of YACs and integration with prokaryotic cloning system
For more detailed mapping and isolation of specific landmarks such as genes
and polymorphisms it is necessary to integrate YACs with prokaryotic cloning
systems containing smaller inserts. Direct screening of cosmid arrays with
YACs, and subcloning of individual YACs into lambda phage and cosmid vectors
will be carried out as part of the course.
3. Assembly of chromosome specific cosmid contigs
Cosmid library screening techniques will be discussed. Filters containing
hundreds of cosmids will be screened with YACs. YAC and cosmid based
techniques will be used to generate cosmid contigs.
4. FISH analysis of YAC and cosmid clones
YACs and cosmids will be mapped on metaphase chromosomes. Selected cosmid
clones will be mapped on extended chromatin fibers for high resolution.
5. Retrofitting and fragmentation of YACs
Manipulation of YAC inserts e.g. the joining of two or more inserts by
recombination between YACs, or the introduction of individual YACs into
heterologous cells for further study of the function of the cloned
sequences, may require alteration of the selectable markers in the YAC.
Methods will be used to retrofit such markers into the vector sequences.
Recombination in yeast can also be used to fragment YACs and generate a
nested set of shorter artificial chromosomes, as well as to amplify the YAC
DNA over the yeast chromosomal background. YAC fragmentation and
amplification experiments will be performed. All these methods will also
provide experience in yeast transformation.
6. Application of YACs to specific problems in human genetics
The particular features of YAC cloning have resulted in the development of
new approaches to solving problems in human genetics. Specific examples will
be presented both as part of the experimental work and in discussion.
7. Gene hunting
Experiments will include cDNA and non cDNA based approaches. cDNA
libraries will be screened with cosmid genomic DNA inserts. In addition
cDNAs will be isolated using cDNA selection techniques. These techniques
will be compared with exon trapping, a technique which does not require
prior knowledge of the expression pattern of a gene.
8. YAC transfer to mammalian cells
YAC transfer to mammalian cells by spheroplast fusion will be undertaken
using a short term assay to determine transfer efficiency.
Informal tutorials and evening lectures (by invited speakers) will
supplement the laboratory sessions.
DUNCAN CAMPBELL (Oxford), ANGELA DAVIES (London), DAVID MARKIE (London),
TONY MONACO (Oxford), IOANNIS RAGOUSSIS (London), CAROL WISE (Texas).
Confirmed speakers include:
ANDREA BALLABIO (Milan) ADRIAN BIRD (Edinburgh), ALAN BUCKLER
(Massachusetts), IAN DUNHAM (Hinxton), MICHAEL LOVETT (Texas), CRAIG
Applicants should be post-doctoral (or close to) scientists engaged in
relevant research. Applicants with molecular biology and/or some cell
culture experience will be particularly welcome. The course is subsidised
by the Wellcome Trust for scientists based in academic institutions anywhere
in the world. This is a residential course, without exception, and there is
a charge of 400 pounds towards board and lodging.
There are no formal application forms, but applicants should send a copy of
their full CV together with a 300 word outline of their research plans to Dr
Pelin Faik, Course Co-ordinator, Division of Biochemistry & Molecular
Biology, UMDS, Guy's Campus, London Bridge SE1 9RT.
Tel: 0171 403 6998 Fax: 0171 407 5281 Email: wss at umds.ac.uk
Closing date for applications is 31st March 1995.
/ \ / | / Alan Young @ uk.ac.ox.vax
/____/ / ___ __ |___/
/ / / / | / | / "The bigger they are,
/ \_/\__\__/|_/ /_________/ the harder they fall on you."
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