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Construction method of porphyra yezoensis plastid genetic transformation vector

A technology of Porphyra zebra and plastid inheritance, applied in the field of bioengineering, to achieve the effects of high expression efficiency, stable pure line of offspring, and good safety

Inactive Publication Date: 2014-03-26
OCEAN UNIV OF CHINA
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Problems solved by technology

There is no report on the genetic transformation of Porphyra plastids

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  • Construction method of porphyra yezoensis plastid genetic transformation vector

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Embodiment Construction

[0024] The technical solutions of the present invention will be further described below in conjunction with the accompanying drawings and specific embodiments.

[0025] A method for constructing a plastid genetic transformation vector of Porphyra zebrais, comprising the following steps::

[0026] (1) Construction of a backbone vector containing rrsB-trnI-trnA-rrsL homologous recombination fragments. ① Cloning of plastid homologous recombination fragments. The rrsB and trnI-trnA-rrsL intergenic regions were selected as the insertion sites of exogenous genes by analyzing the complete plastid genome sequence of P. The following primers were synthesized by Shanghai Yingwei Jieji Biological Co., Ltd., and specific enzyme cutting sites and protective bases were added to the 5' end of each primer.

[0027] XhoI-PyT-F1 5′-CCGCTCGAGGAATCACTGGGCGTAAA-3′

[0028] SacI-PyT-R1 5′-CGAGCTCTTCGCTAATGCTTCAAACTA-3′

[0029] Using Porphyra zebra plastid DNA as a template, it was amplified wi...

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Abstract

The invention discloses a construction method of porphyra yezoensis plastid genetic transformation vector, which comprises the steps of building skeleton vector containing rrsB-trnI-trnA-rrsL homologous recombination segments, building porphyra yezoensis plastid expression vector containing antibiotic screening gene expression cassettes, and building porphyra yezoensis plastid genome targeted integration expressed vector. The method adopts the thought of plastid genetic transformation, and provides a vector basis for plastid genetic transformation, so as to realize the advantages that the expression efficiency of plastid genetic transformation foreign gene is high, gene from pronucleus is not required to be modified, the safety is good, clon is easy to keep, and posterity is not separated.

Description

technical field [0001] The invention belongs to the technical field of bioengineering, and relates to a method for constructing a plastid genetic transformation carrier of Porphyra zebrae. Background technique [0002] Porphyra is a kind of cultivated seaweed with the highest economic value at present. my country is a big porphyra cultivation country, and the annual output of dried porphyra reaches 16,000 tons. In the north of the Yangtze River, it is dominated by Pyropia yezoensis. In the south of the Yangtze River, especially Fujian , Zhejiang mainly cultivates Pyropia haitanensis. In recent years, with the expansion of cultivation area, seaweed cultivation is facing severe situation of species degradation and frequent occurrence of diseases. Therefore, it is one of the important ways to solve the above-mentioned problems by using modern molecular biology methods to carry out genetic engineering research on laver and to cultivate new stress-resistant varieties of laver. ...

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/66C12N15/82
Inventor 孔凡娜刘伟勋茅云翔曹敏
Owner OCEAN UNIV OF CHINA
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