Inactivation of the mouse HPRT locus by a 203-bp retroposon insertion and a 55-kb gene-targeted deletion: establishment of new HPRT-deficient mouse embryonic stem cell lines.
To obtain useful hypoxanthine phosphoribosyl-transferase (HPRT)-deficient mouse ES cell lines, two different methods were employed: (i) selection of spontaneous 6-TG-resistant mutants and (ii) gene targeting of the HPRT locus. The first approach resulted in the establishment of E14.1TG3B1, a spontaneous HPRT-deficient cell line with an insertional mutation of 203 bp in the third exon of the HPRT gene. The insert is highly homologous to the B2 mouse repetitive element and has all the expected retroposon characteristics, thus providing an example of gene inactivation by retroposon insertion. This clone exhibited stable 6-TG resistance and high germ-line transmission frequency. Thus E14.1TG3B1 is a useful ES cell line for modifying the mouse genome using the HPRT gene as a selection marker and for transmission at a high frequency into the mouse germ line. The second approach resulted in a 55-kb deletion of the mouse HPRT locus, demonstrating the feasibility of replacement-targeting vectors to generate large genomic DNA deletions.
Tsuda
H
H
Cell Genesys, Inc., Foster City, California 94404, USA.
Maynard-Currie
C E
CE
Reid
L H
LH
Yoshida
T
T
Edamura
K
K
Maeda
N
N
Smithies
O
O
Jakobovits
A
A
D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D018390:Gene Targeting; D018095:Germ-Line Mutation; D007041:Hypoxanthine Phosphoribosyltransferase / Q000235:genetics*; D051379:Mice; D008969:Molecular Sequence Data; D016254:Mutagenesis, Insertional; D018626:Retroelements; D017384:Sequence Deletion; D013234:Stem Cells
D018626:Retroelements; D007041:Hypoxanthine Phosphoribosyltransferase