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Bombyx mori BmHel-8 transposon with function of enhanser

A transposon, silkworm technology, applied in the direction of transferase, introduction of foreign genetic material using a vector, and cells modified by the introduction of foreign genetic material, etc., can solve the problems of Helitron transposon research stay and so on.

Inactive Publication Date: 2014-07-16
CHONGQING UNIV
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  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] Although Helitron transposons of many species have been reported successively in recent years, the research on Helitron transposons still remains on genome-wide identification
Why do so many Helitron transposon sequences remain in eukaryotic genomes? What function does it have? Still little reported

Method used

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  • Bombyx mori BmHel-8 transposon with function of enhanser

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0019] Identification of the BmHel-8 transposon in the silkworm: Using a combination of structure-based and homology search methods to search for the Helitron transposon sequence in the 9-fold genome sequence of the silkworm. Results The BmHel-8 transposon was identified in the silkworm genome. The consensus sequence of the transposon is 421 bp in length (see SEQ ID NO: 1).

Embodiment 2

[0021] Identification of the enhanced effect of BmHel-8 transposon on downstream gene expression: In order to detect whether BmHel-8 has a regulatory effect on the expression of adjacent genes, we conducted a dual-luciferase reporter gene transfection experiment. We first constructed the A3 promoter sequence of the silkworm to the 5' end of the reporter gene of the firefly luciferase reporter gene vector (pGL3Basic Vector), and then constructed the sequence of BmHel-8 in front of the A3 promoter. Then the constructed vector and the internal reference vector (renilla luciferase reporter gene) were co-transfected into silkworm ovary cells (BmN), and the enzyme activity was measured 48 hours after transfection. It was found that when BmHel-8 was inserted into the 5' end of the luciferase reporter gene, the expression level of the reporter gene was significantly increased (P figure 1 ).

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Abstract

The invention relates to a rolling ring type transposon (BmHel-8) which comes from bombyx mori and has remarkably enhanced downstream gene expression. An enhanser is composed of a nucleotide sequence expressed by SEQ ID No:1. The BmHel-8 transposon is obtained from a bombyx mori genome by means of authentication through the bioinformatic method, and it is verified that the BmHel-8 transposon has the function of the enhanser through the dual-luciferase reporter gene detection method. The BmHel-8 transposon is beneficial to not only acquisition of highly abundant objective protein to promote the development of bombyx mori bioreactors but also the improvement on bombyx mori varieties.

Description

technical field [0001] The field of biotechnology related to the present invention is mainly the field of silkworm transgenic bioreactor and variety improvement. Background technique [0002] Rolling circle transposons (Helitron) were first discovered in the model organisms Arabidopsis, rice and nematode genomes and were classified as DNA transposons (Kapitonov and Jurka2001). However, the sequence structure and transposition mechanism of Helitron transposons are completely different from other DNA transposons. In the transposition mechanism, since the transposase encoded by the complete Helitron transposon contains the conserved motifs of the replication initiation (Rep) and the helicase (Helicase), they are considered to transpose by a rolling circle mechanism. And it is biased to insert into the AT dinucleotide of the genome. In terms of sequence structure, their 5' end starts with TC dinucleotide, and their 3' end ends with CTRR (R stands for A or G) four nucleotides, ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N9/12C12N15/85C12N5/10
CPCC12N9/1241
Inventor 张泽韩民锦
Owner CHONGQING UNIV