Introduction of foreign genes into silkworm eggs by electroporation and its application in transgenic vector test.
Electroporation as a methodology to introduce foreign genes into silkworm eggs was systematically analyzed. The foreign gene in both the newly hatched and 3rd instar larva DNA can be detected by PCR. The amount of foreign gene in 3rd instar larva DNA was about 1/1000 of that in newly hatched larva DNA. The ratio of foreign gene entering into silkworm eggs was voltage dependent and showed significant difference between the tested silkworm strains. When the piggyBac transposon system was applied, the effect of nuclear localization signal (NLS) peptide and the in vitro transcribed transposase mRNA on the transposition rate has been measured. Results showed that the in vitro transcribed transposase mRNA facilitated transposition to take place earlier and NLS could result in higher transposition probability and earlier transposition as well. When linearized vectors containing varied length of flanking homologous sequences around a reporter gene were introduced into silkworm eggs by electroporation, the one with 2.6 kb total arm length gave higher G1 positive ratio than that with total arm length of 1.5 kb and 800 bp.
Guo
Xiu-Yang
XY
Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, the Chinese Academy of Sciences, 320 Yueyang Road, Shanghai 200031, China.
Dong
Liang
L
Wang
Sheng-Peng
SP
Guo
Ting-Qing
TQ
Wang
Jian-Yang
JY
Lu
Chang-De
CD
D000818:Animals; D030801:Animals, Genetically Modified / Q000235:genetics; D012831:Bombyx / Q000235:genetics* / Q000378:metabolism*; D022202:Chromosomes, Artificial, Bacterial / Q000235:genetics; D004251:DNA Transposable Elements / Q000235:genetics; D018274:Electroporation / Q000379:methods*; D020869:Gene Expression Profiling / Q000379:methods; D018014:Gene Transfer Techniques; D005818:Genetic Engineering / Q000379:methods*; D005822:Genetic Vectors; D010063:Ovum / Q000378:metabolism*; D016133:Polymerase Chain Reaction / Q000379:methods; D014170:Transformation, Genetic / Q000235:genetics
D004251:DNA Transposable Elements