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Identification of yeast artificial chromosomes containing the inversion 16 p-arm breakpoint associated with acute
P Liu1, D F Claxton, P Marlton
1Department of Hematology, University of Texas M.D. Anderson Cancer Center, Houston 77030.
Abstract:
We report the cloning of the chromosome 16 p-arm breakpoint involved in inversion 16(p13;q22) associated with subtype of acute myelomonocytic leukemia (AMML) M4Eo. Inter-Alu polymerase chain reaction (PCR) products from a series of interspecific somatic cell hybrids that contain only small portions of the human chromosome 16 p-arm were generated for use as fluorescent in-situ hybridization (FISH) probes. When applied to patient cells, rapid and unambiguous identification of the inversion resulted. Using FISH analysis, cosmid clones associated with the hybrids were identified that bracketed the p-arm breakpoint. A repeat-free fragment of one of these cosmids (35B11) when used as probe on Southern blots from pulsed-field gels identified rearranged macrorestriction fragments in patient DNA. Yeast artificial chromosomes (YACs) were isolated using sequences derived from cosmids flanking 35B11 in a cosmid contig. Of 4 YACs so identified, 3 were shown by FISH to cross the inversion-16 p-arm breakpoint. Therefore, the breakpoint has been molecularly cloned, and identified as being within these 3 YACs. These clones will facilitate the unraveling of the genetic events associated with inversion-16 and are available tools with immediate clinical application.
Insights
Scientists cloned the chromosome 16 p-arm breakpoint in inversion 16, crucial for acute myelomonocytic leukemia (AMML) M4Eo. This breakthrough offers new tools for understanding and diagnosing this leukemia subtype.
Area of Science:
- Molecular Biology
- Genetics
- Oncology
Background:
- Inversion 16(p13;q22) is associated with acute myelomonocytic leukemia (AMML) M4Eo.
- Identifying the specific breakpoint is key to understanding the genetic mechanisms of this leukemia subtype.
Purpose of the Study:
- To clone the chromosome 16 p-arm breakpoint involved in inversion 16.
- To develop molecular tools for diagnosing AMML M4Eo.
Main Methods:
- Utilized interspecific somatic cell hybrids containing portions of the chromosome 16 p-arm.
- Employed Inter-Alu polymerase chain reaction (PCR) to generate probes.
- Applied fluorescent in-situ hybridization (FISH) for breakpoint identification.
- Used cosmid and yeast artificial chromosome (YAC) contigs for breakpoint mapping.
Main Results:
- Successfully identified and cloned the chromosome 16 p-arm breakpoint.
- Developed FISH probes for rapid and unambiguous identification of inversion 16 in patient cells.
- Isolated YAC clones that span the inversion-16 p-arm breakpoint.
Conclusions:
- The chromosome 16 p-arm breakpoint in inversion 16 has been molecularly cloned.
- The cloned breakpoint and associated YACs are valuable tools for investigating the genetic events in AMML M4Eo.
- These findings have immediate clinical applications for leukemia diagnosis and research.