Isolation and characterization of transcribed sequences from a chromosome 16 hn-cDNA library and the physical mapping of genes and transcribed sequences using a high-resolution somatic cell panel of human chromosome 16.
A hn-cDNA (heteronuclear complementary DNA) library was constructed from a mouse/human somatic cell hybrid, CY18, which contains chromosome 16 as the only human chromosome. Hexamer primers constructed from consensus 5' intron splice sequences were used to generate cDNA from the immature unspliced mRNA. The resulting cDNA library was screened with a total human DNA probe to identify potential human clones. Rescreening was necessary, and use of a mouse-derived clone with homology to 7SL RNA proved successful in eliminating the majority of mouse clones. Thirteen clones had open reading frames, and of those, five showed homology to human sequences in GenBank. Two clones had homology to random partially sequenced cDNAs, one clone was likely to be a GRP78 pseudogene, one clone mapped the PHKG2 gene to 16p11.2-16p12.1, and one clone had homology to human S13 ribosomal protein. All clones except the latter were mapped to a high-resolution somatic cell panel. Although isolation of human chromosome 16 genes from this library was successful, it was apparent that cDNA synthesis was initiated at sites other than intron splice sites, presumably by mispairing of the hexamers.
Whitmore
S A
SA
Department of Cytogenetics and Molecular Genetics, Women's and Children's Hospital, Adelaide, Australia.
Apostolou
S
S
Lane
S
S
Nancarrow
J K
JK
Phillips
H A
HA
Richards
R I
RI
Sutherland
G R
GR
Callen
D F
DF
D000818:Animals; D001483:Base Sequence; D015152:Blotting, Northern; D002874:Chromosome Mapping; D002885:Chromosomes, Human, Pair 16; D003001:Cloning, Molecular; D018076:DNA, Complementary; D000091342:Endoplasmic Reticulum Chaperone BiP; D015723:Gene Library; D006801:Humans; D006822:Hybrid Cells; D051379:Mice; D008969:Molecular Sequence Data; D012326:RNA Splicing; D012689:Sequence Homology, Nucleic Acid; D014158:Transcription, Genetic
D018076:DNA, Complementary; D000091342:Endoplasmic Reticulum Chaperone BiP; C000717247:HSPA5 protein, human; C000717248:Hspa5 protein, mouse