Efficient transient expression system based on square pulse electroporation and in vivo luciferase assay of fertilized fish eggs

Ferenc Müller, Z. Lele, László Váradi, László Menczel, László Orbán

Research output: Contribution to journalArticle

48 Citations (Scopus)

Abstract

Electroporation mediated DNA transfer into fish eggs has been improved by using a train of square pulses. Fertilized eggs of African catfish (Clarias gariepinus), zebrafish (Brachydanio rerio) and rosy barb (Barbus conchonius) were dechorionated enzymatically followed by application of pulses. Efficiency of plasmid DNA delivery was significantly increased by applying multiple pulses on dechorionated eggs. Optimization of physical parameters such as field strength, pulse width and pulse numbers resulted in reproducible transient expression in 25-50% of embryos and larvae by using the firefly luciferase and the E. coli β-galactosidase (lacZ) genes both driven by CMV IE1 promoter. Temporal luceferase expression was assayed using both qualitative (sheet film) and quantitative (scintillation counting) methods in developing embryos and fry in vivo. Spatial expression of lacZ was assayed by histochemical staining. A number of embryos revealed foreign gene product also localised in the vegetal pole of the embryo.

Original languageEnglish
Pages (from-to)27-32
Number of pages6
JournalFEBS Letters
Volume324
Issue number1
DOIs
Publication statusPublished - Jun 7 1993

Fingerprint

Electroporation
Zygote
Luciferases
Fish
Assays
Fishes
Embryonic Structures
Genes
Galactosidases
Firefly Luciferases
DNA
Scintillation
Zebrafish
Escherichia coli
Eggs
Poles
Plasmids
Scintillation Counting
Cyprinidae
Catfishes

Keywords

  • Electroporation
  • Firefly luciferase
  • Fish embryo
  • Gene transfer
  • Transient expression
  • β-Galactosidase

ASJC Scopus subject areas

  • Biochemistry
  • Biophysics
  • Molecular Biology

Cite this

Efficient transient expression system based on square pulse electroporation and in vivo luciferase assay of fertilized fish eggs. / Müller, Ferenc; Lele, Z.; Váradi, László; Menczel, László; Orbán, László.

In: FEBS Letters, Vol. 324, No. 1, 07.06.1993, p. 27-32.

Research output: Contribution to journalArticle

Müller, Ferenc ; Lele, Z. ; Váradi, László ; Menczel, László ; Orbán, László. / Efficient transient expression system based on square pulse electroporation and in vivo luciferase assay of fertilized fish eggs. In: FEBS Letters. 1993 ; Vol. 324, No. 1. pp. 27-32.
@article{678c1b736c3c4d029070705893336c29,
title = "Efficient transient expression system based on square pulse electroporation and in vivo luciferase assay of fertilized fish eggs",
abstract = "Electroporation mediated DNA transfer into fish eggs has been improved by using a train of square pulses. Fertilized eggs of African catfish (Clarias gariepinus), zebrafish (Brachydanio rerio) and rosy barb (Barbus conchonius) were dechorionated enzymatically followed by application of pulses. Efficiency of plasmid DNA delivery was significantly increased by applying multiple pulses on dechorionated eggs. Optimization of physical parameters such as field strength, pulse width and pulse numbers resulted in reproducible transient expression in 25-50{\%} of embryos and larvae by using the firefly luciferase and the E. coli β-galactosidase (lacZ) genes both driven by CMV IE1 promoter. Temporal luceferase expression was assayed using both qualitative (sheet film) and quantitative (scintillation counting) methods in developing embryos and fry in vivo. Spatial expression of lacZ was assayed by histochemical staining. A number of embryos revealed foreign gene product also localised in the vegetal pole of the embryo.",
keywords = "Electroporation, Firefly luciferase, Fish embryo, Gene transfer, Transient expression, β-Galactosidase",
author = "Ferenc M{\"u}ller and Z. Lele and L{\'a}szl{\'o} V{\'a}radi and L{\'a}szl{\'o} Menczel and L{\'a}szl{\'o} Orb{\'a}n",
year = "1993",
month = "6",
day = "7",
doi = "10.1016/0014-5793(93)81525-5",
language = "English",
volume = "324",
pages = "27--32",
journal = "FEBS Letters",
issn = "0014-5793",
publisher = "Elsevier",
number = "1",

}

TY - JOUR

T1 - Efficient transient expression system based on square pulse electroporation and in vivo luciferase assay of fertilized fish eggs

AU - Müller, Ferenc

AU - Lele, Z.

AU - Váradi, László

AU - Menczel, László

AU - Orbán, László

PY - 1993/6/7

Y1 - 1993/6/7

N2 - Electroporation mediated DNA transfer into fish eggs has been improved by using a train of square pulses. Fertilized eggs of African catfish (Clarias gariepinus), zebrafish (Brachydanio rerio) and rosy barb (Barbus conchonius) were dechorionated enzymatically followed by application of pulses. Efficiency of plasmid DNA delivery was significantly increased by applying multiple pulses on dechorionated eggs. Optimization of physical parameters such as field strength, pulse width and pulse numbers resulted in reproducible transient expression in 25-50% of embryos and larvae by using the firefly luciferase and the E. coli β-galactosidase (lacZ) genes both driven by CMV IE1 promoter. Temporal luceferase expression was assayed using both qualitative (sheet film) and quantitative (scintillation counting) methods in developing embryos and fry in vivo. Spatial expression of lacZ was assayed by histochemical staining. A number of embryos revealed foreign gene product also localised in the vegetal pole of the embryo.

AB - Electroporation mediated DNA transfer into fish eggs has been improved by using a train of square pulses. Fertilized eggs of African catfish (Clarias gariepinus), zebrafish (Brachydanio rerio) and rosy barb (Barbus conchonius) were dechorionated enzymatically followed by application of pulses. Efficiency of plasmid DNA delivery was significantly increased by applying multiple pulses on dechorionated eggs. Optimization of physical parameters such as field strength, pulse width and pulse numbers resulted in reproducible transient expression in 25-50% of embryos and larvae by using the firefly luciferase and the E. coli β-galactosidase (lacZ) genes both driven by CMV IE1 promoter. Temporal luceferase expression was assayed using both qualitative (sheet film) and quantitative (scintillation counting) methods in developing embryos and fry in vivo. Spatial expression of lacZ was assayed by histochemical staining. A number of embryos revealed foreign gene product also localised in the vegetal pole of the embryo.

KW - Electroporation

KW - Firefly luciferase

KW - Fish embryo

KW - Gene transfer

KW - Transient expression

KW - β-Galactosidase

UR - http://www.scopus.com/inward/record.url?scp=0027154271&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0027154271&partnerID=8YFLogxK

U2 - 10.1016/0014-5793(93)81525-5

DO - 10.1016/0014-5793(93)81525-5

M3 - Article

C2 - 8504855

AN - SCOPUS:0027154271

VL - 324

SP - 27

EP - 32

JO - FEBS Letters

JF - FEBS Letters

SN - 0014-5793

IS - 1

ER -