The goal of this subproject of the proposal is to use yeast artificial chromosome (YAC) vector and pulsed-field gel electrophoresis (PFGE) technology to clone and analyze two large domains of the human genome that are frequently rearranged in human neuroblastoma. We have identified a specific deletion of the short arm of chromosome 1 (at of distal to 1p32) that is characteristic of human neuroblastomas. We are using molecular probes to localize the region that is consistently deleted in neuroblastomas to a band or subband of chromosome 1. Then we will use these probes to screen the YAc library of human DNA to obtain clones from the region of interest. We will develop a restriction map of this region and search for expressed in neuroblastomas. Candidate genes will be cloned and sequenced; and the function and patterns of expression of these genes will be analyzed. Recently, we have demonstrated that the oncogene N-myc is amplified in about one third of primary neuroblastomas. N-myc amplification is associated with advance stages of disease, rapid tumor progression and a poor prognosis in these patients. We plan to make a YAC vector library from a neuroblastoma cell line containing N-myc amplification. We will clone and analyze the structure of the amplified domain in neuroblastomas containing DMs and HSRs. Given the large size of the domain that is consistently amplified in these tumors, it is possible that there are other expressed genes from the domain that confer a selective advantage. We will may the domain and determine if other conserved genes are expressed from this region. Finally, we will identify the ends of the amplified domain, which may represent sites of recombination and provide insights into the mechanism of gene amplification.
| Haimovich, Gal; Zabezhinsky, Dmitry; Haas, Brian et al. (2016) Use of the MS2 aptamer and coat protein for RNA localization in yeast: A response to ""MS2 coat proteins bound to yeast mRNAs block 5' to 3' degradation and trap mRNA decay products: implications for the localization of mRNAs by MS2-MCP system"". RNA 22:660-6 |
