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ShRNA (short hairpin ribonucleic acid) capable of inhibiting expression of buffalo SUV39H1 gene, lentiviral expression vector, construction method of lentiviral expression vector and applications of shRNA

A technology of gene expression and expression vector, applied in DNA/RNA fragmentation, application, genetic engineering, etc., can solve the problems of low natural reproduction rate of buffalo, low population production performance, etc.

Inactive Publication Date: 2015-06-03
GUANGXI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, the natural reproductive rate of buffalo is very low, and the production performance of the population is low. It is urgent to develop and improve a number of high-tech agricultural technologies closely related to livestock genetic improvement to improve the fecundity of buffalo.

Method used

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  • ShRNA (short hairpin ribonucleic acid) capable of inhibiting expression of buffalo SUV39H1 gene, lentiviral expression vector, construction method of lentiviral expression vector and applications of shRNA
  • ShRNA (short hairpin ribonucleic acid) capable of inhibiting expression of buffalo SUV39H1 gene, lentiviral expression vector, construction method of lentiviral expression vector and applications of shRNA
  • ShRNA (short hairpin ribonucleic acid) capable of inhibiting expression of buffalo SUV39H1 gene, lentiviral expression vector, construction method of lentiviral expression vector and applications of shRNA

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Embodiment Construction

[0029] 1. Construction of related expression vectors

[0030] 1. Construction of buffalo SUV39H1 gene expression vector

[0031] Primers were designed according to the bovine SUV39H1 gene mRNA [NM_001046264.2] sequence published by NCBI GenBank, and XhoI and EcoRI restriction sites were introduced at the 5' ends of the upstream and downstream primers respectively. The primers were:

[0032] SUV39H1-upstream 5'-CTCGAGGGAAAGATGGCGGAAAAT-3' (SEQ.ID.No.7 of the sequence listing),

[0033] SUV39H1-Downstream 5'-GAATTCCCGAAGAGGTATTTGCGGCAG-3' (Sequence Listing SEQ.ID.No.8).

[0034] Fresh buffalo ovarian tissue was taken from the slaughterhouse and put into a 2ml EP tube pre-added with 1ml RNA protection agent. Total RNA was extracted by Trizol lysis and reverse transcribed into cDNA. Using the buffalo ovary cDNA as a template, the full-length ORF sequence of the buffalo SUV39H1 gene (SEQ.ID.No.9) was amplified to obtain the full-length ORF sequence of the buffalo SUV39H1 gene. ...

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Abstract

The invention discloses a shRNA (short hairpin ribonucleic acid) capable of inhibiting the expression of a buffalo SUV39H1 gene. The shRNA is shSUV39H1-259 or shSUV39H1-597 which comprises siRNA (small interfering ribonucleic acid) sense strand of 19nt, loop ring of 9nt, siRNA antisense strand of 19nt, and a termination signal. The shRNA can efficiently inhibit the expression of the buffalo SUV39H1 gene. The invention further comprises a lentiviral expression vector of the shRNA and a construction method of the lentiviral expression vector. Proved by experiments, the expression of the buffalo SUV39H1 gene can be effectively and stably inhibited by applying the shRNA, and in gene function research and the transgenic animal preparation processes, the influence and action mechanism on the expression efficiency of a foreign target transgene can be researched through inhibiting the expression of the SUV39H1 gene, thus providing a new conception and method for the search for improving the expression efficiency of the foreign gene in the transgenic animal.

Description

technical field [0001] The invention relates to a buffalo mottled suppressor homologue 1 (SUV39H1) gene, in particular to a shRNA, a lentiviral expression vector for inhibiting the expression of the buffalo SUV39H1 gene, a construction method and application thereof. Background technique [0002] Buffalo is one of the main large livestock in southern my country. It has biological characteristics such as high temperature and high humidity resistance, strong disease resistance, and resistance to rough feeding. It is very suitable for breeding in rural areas in southern my country. Buffalo milk has good quality and high nutritional value. Its dry matter content, milk fat percentage and total protein content are twice that of ordinary milk, especially the content of casein (4.14%), which is 16 times that of black-and-white cows ( 0.26%), it is a high-quality raw material for making high-grade cheese. Therefore, it is the development trend of the buffalo animal husbandry industr...

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/113C12N15/867
Inventor 李湘萍石德顺林浪
Owner GUANGXI UNIV