REPSA: A method for determining the sequence specificity of DNA binding ligands

We developed a novel combinatorial method termed restriction endonuclease protection selection and amplification (REPSA) to identify consensus binding sites of DNA-binding ligands. REPSA uses a unique enzymatic selection based on the inhibition of cleavage by a type IIS restriction endonuclease, an enzyme that cleaves DNA at a site distal from its recognition sequence. Sequences bound by a ligand are protected from cleavage while unprotected sequences are cleaved. This enzymatic selection occurs in solution under mild conditions and is dependant only on the DNA-binding ability of the ligand. Thus, REPSA is useful for a broad range of ligands including all classes of DNA-binding ligands, weakly binding ligands, mixed populations of ligands, and unknown ligands. Here I describe REPSA and the application of this method to select the consensus DNA-binding sequences of three representative DNA-binding ligands; a nucleic acid (triplex-forming single-stranded DNA), a protein (the TATA-binding protein), and a small molecule (Distamycin A). These studies generated new information regarding the specificity of these ligands in addition to establishing their DNA-binding sequences. ^

Cost analysis of opt-in consent procedures for the Texas statewide immunization information system, ImmTrac

Background. Childhood immunization programs have dramatically reduced the morbidity and mortality associated with vaccine-preventable diseases. Proper documentation of immunizations that have been administered is essential to prevent duplicate immunization of children. To help improve documentation, immunization information systems (IISs) have been developed. IISs are comprehensive repositories of immunization information for children residing within a geographic region. The two models for participation in an IIS are voluntary inclusion, or "opt-in," and voluntary exclusion, or "opt-out." In an opt-in system, consent must be obtained for each participant, conversely, in an opt-out IIS, all children are included unless procedures to exclude the child are completed. Consent requirements for participation vary by state; the Texas IIS, ImmTrac, is an opt-in system.^ Objectives. The specific objectives are to: (1) Evaluate the variance among the time and costs associated with collecting ImmTrac consent at public and private birthing hospitals in the Greater Houston area; (2) Estimate the total costs associated with collecting ImmTrac consent at selected public and private birthing hospitals in the Greater Houston area; (3) Describe the alternative opt-out process for collecting ImmTrac consent at birth and discuss the associated cost savings relative to an opt-in system.^ Methods. Existing time-motion studies (n=281) conducted between October, 2006 and August, 2007 at 8 birthing hospitals in the Greater Houston area were used to assess the time and costs associated with obtaining ImmTrac consent at birth. All data analyzed are deidentified and contain no personal information. Variations in time and costs at each location were assessed and total costs per child and costs per year were estimated. The cost of an alternative opt-out system was also calculated.^ Results. The median time required by birth registrars to complete consent procedures varied from 72-285 seconds per child. The annual costs associated with obtaining consent for 388,285 newborns in ImmTrac's opt-in consent process were estimated at $702,000. The corresponding costs of the proposed opt-out system were estimated to total $194,000 per year. ^ Conclusions. Substantial variation in the time and costs associated with completion of ImmTrac consent procedures were observed. Changing to an opt-out system for participation could represent significant cost savings. ^