Rop small GTPases are plant-specific signaling proteins with roles in pollen and vegetative cell growth, abscisic acid signal transduction, stress responses, and pathogen resistance. We have characterized the rop family in the monocots maize (Zea mays) and rice (Oryza sativa). The maize genome contains at least nine expressed rops, and the fully sequenced rice genome has seven. Based on phylogenetic analyses of all available Rops, the family can be subdivided into four groups that predate the divergence of monocots and dicots; at least three have been maintained in both lineages. However, the Rop family has evolved differently in the two lineages, with each exhibiting apparent expansion in different groups. These analyses, together with genetic mapping and identification of conserved non-coding sequences, predict orthology for specific rice and maize rops. We also identified consensus protein sequence elements specific to each Rop group. A survey of ROP-mRNA expression in maize, based on multiplex reverse transcriptase-polymerase chain reaction and a massively parallel signature sequencing database, showed significant spatial and temporal overlap of the nine transcripts, with high levels of all nine in tissues in which cells are actively dividing and expanding. However, only a subset of rops was highly expressed in mature leaves and pollen. Intriguingly, the grouping of maize rops based on hierarchical clustering of expression profiles was remarkably similar to that obtained by phylogenetic analysis. We hypothesize that the Rop groups represent classes with distinct functions, which are specified by the unique protein sequence elements in each group and by their distinct expression patterns.Rho family GTPases are well-characterized regulators of cellular morphogenesis in fungal, insect, and mammalian cells (Lu and Settleman, 1999;Hall and Nobes, 2000;Settleman, 2001). A plant-specific family of Rho homologs, known as the Rop family (Rhorelated protein from plants), has important roles in plant development Yang, 2002). Rops have been linked to the regulation of pollen tube and root hair growth, vegetative cell expansion, cell wall synthesis, and cell proliferation in the meristem (Valster et al., 2000;Zheng and Yang, 2000;. They carry out at least some of their developmental functions through F-actin (Kost et al., 1999a;Fu et al., , 2002, a crucial component in plant cell morphogenesis (Fowler and Quatrano, 1997;Kost et al., 1999b). In addition to roles in development, Rops may have significant roles in signaling pathways through which plants respond to their environment. For example, Rop plays a role in abscisic acid signaling (Lemichez et al., 2001;Zheng et al., 2002) and in tolerance to oxygen deprivation (Baxter-Burrell et al., 2002). Furthermore, overexpression of a constitutively active form of the rice (Oryza sativa) Rop OsRac1 promotes both the generation of reactive oxygen species and a pathogen-induced cell death response (Kawasaki et al., 1999;Ono et al., 2001). Similarly, maize (Zea mays) ROPs can in...

We have designed, tested, and validated synthetic DNA molecules that may be used as reference standard controls in the simultaneous detection of mutations in one or more genes. These controls consist of a mixture of oligonucleotides (100 to 120 bases long) each designed for the detection of one or more disease-causing mutation(s), depending on the proximity of the mutations to one another. Each control molecule is identical to 80 to 100 bases that span the targeted mutations. In addition, each oligonucleotide is tagged at the 5' and 3' ends with distinct nucleic acid sequences that allow for the design of complementary primers for polymerase chain reaction amplification. We designed the tags to amplify control molecules comprising 32 CFTR mutations, including the American College of Medical Genetics minimum carrier screening panel of 23, with one pair of primers in a single tube. We tested the performance of these controls on many platforms including the Applied Biosystems/Celera oligonucleotide ligation assay and the Tm Bioscience Tag-It platforms. All 32 mutations were detected consistently. This simple methodology allows for maximum flexibility and rapid implementation. It has not escaped our notice that the design of these molecules makes possible the production of similar controls for virtually any mutation or sequence of interest.

Synthetic quality control materials proved to be valuable in troubleshooting underperforming assays and complement existing genomic controls. The study also revealed a strong need for increased quality control in the postanalytical phase of testing.