Isolation of murine telomere-proximal sequences by affinity capture and PCR

Journal Article · Tue Oct 10 04:00:00 EDT 1995 · Genomics

DOI:https://doi.org/10.1006/geno.1995.9958· OSTI ID:443858

Rounds, D; Ward, D C; Brueckner, M ^[1]

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1. Yale Univ. School of Medicine, New Haven, CT (United States)

We describe a method of selectively enriching for murine telomere-proximal sequences using affinity capture followed by PCR amplification. The telomeric fragments were selected from NotI-digested and lambda exonuclease-resected mouse genomic DNA by annealing to a biotinylated riboprobe containing multiple copies of the telomere repeat (TTAGGG){sub n}. The resultant DNA-RNA hybrids were selectively retained on a matrix with covalently bound avidin. The captured DNA was then specifically released by ribonuclease action, and PCR amplification was performed using mouse repeat primers. The PCR products were cloned and used to screen a mouse genomic cosmid library, and the resultant cosmid clones were analyzed by fluorescence in situ hybridization. Ten of 70 clones analyzed gave telomere-proximal hybridization signals, indicating an at least 500-fold enrichment for telomere-proximal sequences. 24 refs., 4 figs., 1 tab.

• Sponsoring Organization: USDOE
• OSTI ID: 443858
• Journal Information: Genomics, Journal Name: Genomics Journal Issue: 3 Vol. 29; ISSN 0888-7543; ISSN GNMCEP
• Country of Publication: United States
• Language: English

Related Subjects

44 INSTRUMENTATION
INCLUDING NUCLEAR AND PARTICLE DETECTORS
55 BIOLOGY AND MEDICINE
BASIC STUDIES
BIOASSAY
BIOLOGICAL MARKERS
CHROMOSOMES
COSMIDS
DNA
DNA HYBRIDIZATION
DNA SEQUENCING
DNA-CLONING
EVALUATION
FLUORESCENCE
GENES
GENETIC MAPPING
MICE
POLYMERASE CHAIN REACTION
SIZE
STRUCTURE-ACTIVITY RELATIONSHIPS
TELOMERES