Human ribosomal RNA gene cluster: identification of the proximal end containing a novel tandem repeat sequence.
Human ribosomal RNA genes (rDNA) are arranged as tandem repeat clusters on the short arms of five pairs of acrocentric chromosomes. We have demonstrated that a majority of the rDNA clusters are detected as 3-Mb DNA fragments when released from human genomic DNA by EcoRV digestion. This indicated the absence of the EcoRV restriction site within the rDNA clusters. We then screened for rDNA-positive cosmid clones using a chromosome 22-specific cosmid library that was constructed from MboI partial digests of the flow-sorted chromosomes. Three hundred twenty rDNA-positive clones negative for the previously reported distal flanking sequence (pACR1) were chosen and subjected to EcoRV digestion. Seven clones susceptible to EcoRV were further characterized as candidate clones that might have been derived from the junctions of the 3-Mb rDNA cluster. We identified one clone containing part of the rDNA unit sequence and a novel flanking sequence. Detailed analysis of this unique clone revealed that the coding region of the last rRNA gene located at the proximal end of the cluster is interrupted with a novel sequence of approximately 147 bp that is tandemly repeated and is connected with an intervening 68-bp unique sequence. This junction sequence was readily amplified from chromosomes 21 and 15 as well as 22 using the polymerase chain reaction. Fluorescence in situ hybridization further indicated that the approximately 147-bp sequence repeat is commonly distributed among all the acrocentric short arms.
Sakai
K
K
Department of Molecular Biology, Keio University School of Medicine, Tokyo, Japan.
Ohta
T
T
Minoshima
S
S
Kudoh
J
J
Wang
Y
Y
de Jong
P J
PJ
Shimizu
N
N
D000818:Animals; D001483:Base Sequence; D016466:CHO Cells; D002892:Chromosomes, Human, Pair 22; D003001:Cloning, Molecular; D006224:Cricetinae; D004275:DNA, Ribosomal; D016521:Electrophoresis, Gel, Pulsed-Field; D006801:Humans; D006822:Hybrid Cells; D008969:Molecular Sequence Data; D005810:Multigene Family; D012335:RNA, Ribosomal / Q000235:genetics*; D012091:Repetitive Sequences, Nucleic Acid
D004275:DNA, Ribosomal; D012335:RNA, Ribosomal