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<h2><center>Photosynthetic Mutant Library
(PML)</center></h2>
<h4>A genetic resource in maize that is tailored for studies
of chloroplast biogenesis</h4>
<p>PML is a library of Mu- induced non-photosynthetic maize
mutants, corresponding DNA samples, and phenotypic data.
Mutants contributed to PML exhibit either a seedling
chlorophyll deficiency (e.g. pale green, yellow, virescent,
albino, or striated leaves) or a high chlorophyll
fluorescent (hcf) phenotype. Prior studies indicate that a
disruption of most any step in the biogenesis of the
chloroplast (i.e. chloroplast gene expression, lipid or
pigment synthesis, light perception, membrane assembly,
protein import) results in one of the phenotypes contributed
to PML. The collection currently consists of ~1700
independently-arising mutants. It is being expanded to ~2500
mutants, at which point we anticipate the screen will be
saturated.</p>
<p>Researchers can use this resource in two distinct ways.
The pooled DNA samples can be used as a "reverse genetic
resource" to identify mutants with Mu insertions in genes of
known sequence with a postulated role in chloroplast
biogenesis. Alternatively, the phenotype database (a catalog
of pigment deficiency, photosynthetic enzyme content and
chloroplast RNA defects) can be searched for phenotypes of
interest, and corresponding seed requested.</p>
<h4>PML as a source of mutant alleles of genes of known
sequence</h4>
<p>The PML collection is anticipated to include mutations in
the vast majority of genes that play non-redundant roles in
the establishment of a photosynthetically-competent
chloroplast. The relatively small number of DNA samples can
be screened by PCR in a cost-effective manner for Mu
insertions into genes of known sequence with proposed roles
in chloroplast biogenesis. It can be used to obtain the
first mutant alleles in genes of known sequence, or to
provide additional alleles of genes for which one mutant
allele is already available.</p>
<h4>Which reverse genetic resource is best for my
project?</h4>
<p>Several resources related to PML are now available to all
academic researchers: maize <a href="http://mtm.cshl.org/" target="_blank">MTM
Mutator lines</a>, <a href="http://www.biotech.wisc.edu/Arabidopsis/" target="_blank">Arabidopsis
T-DNA populations</a> and transposon lines in <a href="http://spot.cshl.org/genetrap_database/mainframe.html" target="_blank">Arabidopsis</a>
and <a href="http://gremlin3.zool.iastate.edu/" target="_blank">maize</a>
for which sequence tags flanking transposon-insertions are
being generated. Each has different attributes which suits
it for distinct purposes.</p>
<h4>Choice #1: Maize vs Arabidopsis</h4>
<blockquote><b>Factors to consider:</b>
<p>The large maize seed supports the robust growth of
non-photosynthetic seedlings (see adjacent <a href="javascript:newwindow()">image</a>
of 10 day old albino seedling (~0.5gm leaf tissue)
growing happily in soil ), facilitating biochemical
analyses of mutant leaf tissue. Although
non-photosynthetic Arabidopsis mutants can be grown on
sterile sugar medium, this is costly and slow, and alters
the physiology of the plant in ways that may confound the
interpretation of mutant phenotypes.</p>
<p>Arabidopsis is much more easily transformed.
Arabidopsis mutants are best suited for approaches that
require the frequent generation of transgenic plants.</p>
<p>Mutations that cause severe and global defects in
chloroplast gene expression (e.g. the complete loss of
plastid ribosomes) can be recovered in maize but may not
be recoverable in Arabidopsis due to the presence of
genes in the Arabidopsis chloroplast genome that are
required for cell viability.</p>
<p>The availability of the entire Arabidopsis genome
sequence offers opportunities that are not yet available
in maize. However, it is often possible to predict
orthologous relationships between Arabidopsis genes and
maize EST sequences, simplifying movement between the two
organisms.</p>
<blockquote>Bottom line: a combined approach will often
be the best!</blockquote></blockquote>
<p><b>Choice #2: MTM vs PML vs RescueMu.</b></p>
<blockquote>Advantages of <a href="http://mtm.cshl.org/" target="_blank">MTM</a>
and <a href="http://gremlin3.zool.iastate.edu/zmdb/RescueMu.html" target="_blank">RescueMu</a>
sequence tags: no presupposition about mutant phenotype,
therefore hits may be recovered in genes whose disruption
results in no obvious mutant phenotype or in an embryo
lethal phenotype. Such mutants will not be recovered in
PML.
<p>RescueMu sequence tag screening has the advantage that
screens are done <i>in silico</i>. However, the number of
such sequence tags is currently rather limited.</p>
<p>For both these resources, as for PML, users need to
establish whether hits detected represent germinal or
somatic insertions, and whether they are
genetically-linked to any mutant phenotype detected..</p>
<p>If a mutation in your gene does result in one of the
phenotypes in the PML collection, it will be advantageous
to screen PML. Less time will be spent sorting through
useless alleles in which the Mu insertion does not
disrupt gene function.</p>
<p>Bottom line: If you are confident that your protein is
chloroplast-localized and have good reason to believe
that it plays a role in chloroplast gene expression or
the assembly or functioning of the photosynthetic
apparatus, PML is probably the best place to start. It
doesn't cost much and may get you useful mutants more
easily than MTM. If PML comes up empty, try MTM!</p></blockquote>
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<a href="http://pml.uoregon.edu" target="_top">Home</a></center></h2>
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