DETERMINATION OF TRANSIENT OR STABLE NEO EXPRESSION LEVELS IN MAMMALIAN-CELLS

被引：12

作者：

DUCH, M ^{[1
]}

PALUDAN, K ^{[1
]}

PEDERSEN, L ^{[1
]}

JORGENSEN, P ^{[1
]}

KJELDGAARD, NO ^{[1
]}

PEDERSEN, FS ^{[1
]}

机构：

[1] AARHUS UNIV,DEPT MOLEC BIOL & PLANT PHYSIOL,CE MOLLERS ALLE 130,DK-8000 AARHUS,DENMARK

来源：

GENE | 1990年 / 95卷 / 02期

基金：

欧盟地平线“2020”;

关键词：

co-transfection; densitometry; ELISA; Geneticin; marker gene; neomycin phosphotransferase; protein measurement; Recombinant DNA;

D O I：

10.1016/0378-1119(90)90373-Y

中图分类号：

Q3 [遗传学];

学科分类号：

071007 ; 090102 ;

摘要：

We report an extension of the neomycin phosphotransferase II dot-blot assay to allow more rapid and sensitive quantitative determination of the neo gene product in crude mammalian cell extracts. Our procedure, based upon the dot-blot assay of Platt and Yang [Anal. Biochem. 162 (1987) 502-514], measures both the enzymatic activity and the protein content of a cell extract by scanning with an enzyme-linked immunosorbent assay reader, using the same sample rather than parallel samples for both measurements. We show this assay to be comparable to the chloramphenicol acetyltransferase assay in sensitivity. Therefore, apart from being a useful selectable marker gene, the neo gene is a convenient reporter gene in studies of stable, as well as transient, expression. © 1990.

引用

页码：285 / 288

页数：4

共 14 条

[1] NUCLEOTIDE-SEQUENCE AND EXACT LOCALIZATION OF THE NEOMYCIN PHOSPHOTRANSFERASE GENE FROM TRANSPOSON TN5
BECK, E
LUDWIG, G
AUERSWALD, EA
REISS, B
SCHALLER, H
[J]. GENE, 1982, 19 (03) : 327 - 336
[2] A NEW DOMINANT HYBRID SELECTIVE MARKER FOR HIGHER EUKARYOTIC CELLS
COLBEREGARAPIN, F
HORODNICEANU, F
KOURILSKY, P
GARAPIN, AC
[J]. JOURNAL OF MOLECULAR BIOLOGY, 1981, 150 (01) : 1 - 14
[3] Gorman C., 1985, DNA CLONING PRACTICA, V1, P143
[4] RECOMBINANT GENOMES WHICH EXPRESS CHLORAMPHENICOL ACETYLTRANSFERASE IN MAMMALIAN-CELLS
GORMAN, CM
MOFFAT, LF
HOWARD, BH
[J]. MOLECULAR AND CELLULAR BIOLOGY, 1982, 2 (09) : 1044 - 1051
[5] Haas M J, 1975, Methods Enzymol, V43, P611
[6] MAMMALIAN EXPRESSION-AND-TRANSMISSION VECTOR DERIVED FROM AKV MURINE LEUKEMIA-VIRUS
JENSEN, NA
JORGENSEN, P
KJELDGAARD, NO
PEDERSEN, FS
[J]. GENE, 1986, 41 (01) : 59 - 65
[7] MCGRATH MS, 1980, CONT TOPICS IMMUNOLO, P157
[8] DIFFERENT RELATIVE EXPRESSION FROM 2 MURINE LEUKEMIA-VIRUS LONG TERMINAL REPEATS IN UNINTEGRATED TRANSFECTED DNA AND IN INTEGRATED RETROVIRAL VECTOR PROVIRUSES
PALUDAN, K
DAI, HY
DUCH, M
JORGENSEN, P
KJELDGAARD, NO
PEDERSEN, FS
[J]. JOURNAL OF VIROLOGY, 1989, 63 (12) : 5201 - 5207
[9] PALUDAN K, 1989, GENE, V85, P423
[10] DOT ASSAY FOR NEOMYCIN PHOSPHOTRANSFERASE ACTIVITY IN CRUDE CELL-EXTRACTS
PLATT, SG
YANG, NS
[J]. ANALYTICAL BIOCHEMISTRY, 1987, 162 (02) : 529 - 535

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