Introduction of Foreign Genes into Silkworm Eggs by Electroporation and Its Application in Transgenic Vector Test  

作　　者：Xiu-YangGUO LiangDONG Sheng-PengWANG Ting-QingGUO Jian-YangWANG Chang-DeLU 

机构地区：[1]InstituteofBiochemistryandCellBiology,ShanghaiInstitutesforBiologicalSciences,theChineseAcademyofSciences320YueyangRoad,Shanghai200031,China

出　　处：《Acta Biochimica et Biophysica Sinica》2004年第5期323-330,共8页生物化学与生物物理学报（英文版）

基　　金：This work was supported by the grants from the Natural Science Foundation of China (No. 30370326) and the Science and Technology Commit-tee of Shanghai (No. 024319209).

摘　　要：Electroporation as a methodology to introduce foreign genes into silkworm eggs was system- atically 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 transposi- tion rate has been measured. Results showed that the in vitro transcribed transposase mRNA facilitated trans- position 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 re- porter 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.Electroporation as a methodology to introduce foreign genes into silkworm eggs was system- atically 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 transposi- tion rate has been measured. Results showed that the in vitro transcribed transposase mRNA facilitated trans- position 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 re- porter 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.

关 键 词：silkworm eggs gene introduction ELECTROPORATION TRANSPOSON homologous sequences 

分 类 号：Q78[生物学—分子生物学]