To solve this problem, we propose a "Phenotype Sequencing" approach in which genes causing the phenotype can be identified directly from sequencing of multiple independent mutants.
Several of these homology-independent duplications were identified in multiple independent mutants, suggesting that homology-independent duplications can be formed in a site-specific manner.
However, owing to the lack of sexual reproduction in this species, the presence of additional mutations might affect growth rate, suggesting that selection should include evaluation of multiple independent mutants for each desired phenotype.
However, if we can obtain multiple independent mutant strains from our phenotype screen, the statistics of independent selection events will quickly help distinguish the true target genes.
For such organisms, phenotype sequencing can open up a fast path for directly identifying a phenotype's genetic causes, for any phenotype where a good screen exists for generating multiple independent mutant strains.
We describe the isolation of multiple, independent mutant lines, including two (vt21 and vt22 ) that possess novel alleles of DAT-1.
Analysis of multiple independent disruption mutant strains for each candidate vic allele failed to find any obvious changes in growth, colony morphology, sporulation, or virulence (data not shown).
In order to have sufficient neurospheres of similar sizes for assay, subsequent experiments used neurospheres after secondary expansion and derived from multiple independent control and mutant embryos.
This nodulation phenotype was observed in the SACPD-C deletion lines A6, MM106 (data not shown) and FN8-10, as well as multiple, independent the point mutant lines; i.e. 194D (V211E), KK24 (C298Δ) and FAM94-41 (D126N).
Mutations in erosion had unique sequences when compared with the paired non-erosion mutations from the same patient, which may not be surprising given the results of microdissection studies demonstrating multiple independent islands of mutant cells rather than diffuse monoclonal expansion [ 9].
This model addresses the question of whether or not the changes in nuclear transport can be explained by the existence of multiple, independent NLSs, assuming transferability between mutant experiments, or on the requirement for cooperativity.
