A simple method for the rapid purification of copia virus-like particles from Drosophila Schneider 2 cells

被引:20
作者
Bachmann, AS
Corpuz, G
Hareld, WP
Wang, G
Coller, BA
机构
[1] Univ Hawaii, Canc Res Ctr Hawaii, Honolulu, HI 96813 USA
[2] GlaxoSmithKline Biol, B-1330 Rixensart, Belgium
[3] Hawaii Biotech Inc, Aiea, HI 96701 USA
关键词
Copia; Drosophila Schneider 2 cells; purification; virus-like particles;
D O I
10.1016/j.jviromet.2003.09.025
中图分类号
Q5 [生物化学];
学科分类号
071010 ; 081704 ;
摘要
Drosophila retrotransposon copia produces virus-like particles (VLPs) in cultured Drosophila cells. The VLPs contain copia RNA and reverse transcriptase activity, and thus, play a major role in copia replication. Here, we report a rapid and simple method for the purification of copia VLPs from Drosophila Schneider 2 (S2) cells. The VLP purification procedure consists of a series of short centrifugation steps and eliminates the tedious and time-consuming performance of sucrose, metrizamide and cesium chloride (CsCl) gradients. The purity and presence of VLPs at different purification steps was monitored by transmission electron microscopy (TEM), immunochemical detection methods, and by Northern blot analysis. In addition to providing a fast protocol for VLP purification, our results also show that copia particles are mainly located in the nucleus of S2 cells. This new protocol may find broad applications for the purification of various other VLPs, and thus, may be of great value to other investigators with similar interests. (C) 2003 Elsevier B.V. All rights reserved.
引用
收藏
页码:159 / 165
页数:7
相关论文
共 24 条
[1]   EVIDENCE FOR TRANSPOSITION OF DISPERSED REPETITIVE DNA FAMILIES IN YEAST [J].
CAMERON, JR ;
LOH, EY ;
DAVIS, RW .
CELL, 1979, 16 (04) :739-751
[2]   THE NUCLEOTIDE-SEQUENCES OF COPIA AND COPIA-RELATED RNA IN DROSOPHILA VIRUS-LIKE PARTICLES [J].
EMORI, Y ;
SHIBA, T ;
KANAYA, S ;
INOUYE, S ;
YUKI, S ;
SAIGO, K .
NATURE, 1985, 315 (6022) :773-776
[3]   EXTRACHROMOSOMAL CIRCULAR COPIES OF THE EUKARYOTIC TRANSPOSABLE ELEMENT COPIA IN CULTURED DROSOPHILA CELLS [J].
FLAVELL, AJ ;
ISHHOROWICZ, D .
NATURE, 1981, 292 (5824) :591-595
[4]  
GATEFF E, 1978, SCIENCE, V200, P1448, DOI 10.1126/science.96525
[5]   PURIFICATION AND PARTIAL CHARACTERIZATION OF VIRUS-LIKE PARTICLES FROM SCHNEIDER LINE-2 DROSOPHILA CELLS [J].
GROSS, RH ;
HORMANN, RE ;
SAXE, JC .
ARCHIVES OF BIOCHEMISTRY AND BIOPHYSICS, 1981, 207 (02) :455-459
[6]   INSERTION OF A MOVABLE GENETIC ELEMENT, 297, INTO THE T-A-T-A BOX FOR THE H-3 HISTONE GENE IN DROSOPHILA-MELANOGASTER [J].
IKENAGA, H ;
SAIGO, K .
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA-BIOLOGICAL SCIENCES, 1982, 79 (13) :4143-4147
[7]   Evolution of the copia retrotransposon in the Drosophila melanogaster species subgroup [J].
Jordan, IK ;
McDonald, JF .
MOLECULAR BIOLOGY AND EVOLUTION, 1998, 15 (09) :1160-1171
[8]   VARIATION IN NUMBER OF LONG, OPEN READING FRAMES AMONG THE MEMBERS OF COPIA, A DROSOPHILA RETROTRANSPOSON, AND CDNAS OF COPIA-RELATED RNA IN VIRUS-LIKE PARTICLES [J].
KANDA, T ;
HATTORI, K ;
SAIGO, K .
NUCLEIC ACIDS RESEARCH, 1986, 14 (10) :4369-4369
[9]   RETROVIRUSES IN INVERTEBRATES - THE GYPSY RETROTRANSPOSON IS APPARENTLY AN INFECTIOUS RETROVIRUS OF DROSOPHILA-MELANOGASTER [J].
KIM, A ;
TERZIAN, C ;
SANTAMARIA, P ;
PELISSON, A ;
PRUDHOMME, N ;
BUCHETON, A .
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, 1994, 91 (04) :1285-1289
[10]   CLEAVAGE OF STRUCTURAL PROTEINS DURING ASSEMBLY OF HEAD OF BACTERIOPHAGE-T4 [J].
LAEMMLI, UK .
NATURE, 1970, 227 (5259) :680-+