Chlamydomonas reinhardtii TCTN1 expression plasmid as well as construction method and application thereof

A technology of TCTN1 and expression plasmids, which is applied in the field of bioengineering and can solve problems affecting drug development and formulation of treatment plans

Active Publication Date: 2021-04-16
XUZHOU NORMAL UNIVERSITY
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  • Abstract
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Problems solved by technology

However, the pathogenic molecular mechanism behind it is not well understood, which affects the development of its drugs and the formulation of treatment plans.

Method used

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  • Chlamydomonas reinhardtii TCTN1 expression plasmid as well as construction method and application thereof
  • Chlamydomonas reinhardtii TCTN1 expression plasmid as well as construction method and application thereof
  • Chlamydomonas reinhardtii TCTN1 expression plasmid as well as construction method and application thereof

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[0026] The present invention will be described in further detail below in conjunction with the accompanying drawings and specific embodiments. In the following methods, unless otherwise specified, they are conventional methods.

[0027] A Chlamydomonas reinhardtii TCTN1 expression plasmid comprising a hygromycin B selection gene, an endogenous promoter expressing the TCTN1 gene, TCTN1 gDNA, a 3' terminal 3×HA tag and a termination of TCTN1 transcription The terminator rbcS2; wherein, the nucleotide sequence of the hygromycin B screening gene is shown in SEQ ID.No.11, and the nucleotide sequence of the endogenous promoter expressing the TCTN1 gene is shown in SEQ ID.No.12 The nucleotide sequence of TCTN1 gDNA is shown in SEQID.No.13, the nucleotide sequence of the 3'-end 3×HA tag is shown in SEQ ID.No.14, and the nucleotide sequence of the terminator rbcS2 is shown in As shown in SEQ ID.No.15, the nucleotide sequence of the Chlamydomonas reinhardtii TCTN1 expression plasmid is...

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Abstract

The invention discloses a chlamydomonas reinhardtii TCTN1 expression plasmid as well as a construction method and application thereof. The nucleotide sequence of the chlamydomonas reinhardtii TCTN1 expression plasmid is as shown in SEQ ID No.1. The chlamydomonas reinhardtii TCTN1 expression plasmid comprises a hygromycin B screening gene, an endogenous promoter for expressing a TCTN1 gene, TCTN1 gDNA, a 3 * HA tag at the 3'terminal and a terminator rbcS2 for terminating TCTN1 transcription; The construction method comprises the following steps: cloning a chlamydomonas reinhardtii TCTN1 gene segment; constructing a T vector plasmid containing a TCTN1 fragment; obtaining a terminal phosphorylated TCTN1 fragment; preparing a pHyg-3HA vector; and constructing and verifying the plasmid of the pHyg-TCTN1-3HA. According to the expression plasmid of the chlamydomonas TCTN1, algal strains capable of stably expressing the TCTN1-HA fusion protein can be efficiently screened, so that a chlamydomonas cilia transition region is marked, and the structure of the transition region and the function of a complex are known.

Description

technical field [0001] The invention belongs to the field of bioengineering, and in particular relates to a Chlamydomonas reinhardtii TCTN1 expression plasmid, a construction method and application thereof. Background technique [0002] Chlamydomonas reinhardtii is a single-celled eukaryote with the title of photosynthetic yeast, and it is also one of the model organisms. Its growth conditions are simple, easy to culture, and low in culture costs; the cell cycle is short and its reproductive capacity is strong; the whole genome sequencing of Chlamydomonas has been completed, and the cDNA library has been established; it has cilia, and its cilia structural components are very similar to those of higher animals , synchronous assembly of cilia, easy to observe, separate, extract and purify; the transgenic method of Chlamydomonas is mature. Therefore, Chlamydomonas is widely used in the field of molecular biology as a research model for cilia disease research, plant photosynthe...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/79C12N15/65C12N15/66C12N15/62C07K19/00
Inventor 王亮温馨李春红周慧琳李宇涵程一非王雯
Owner XUZHOU NORMAL UNIVERSITY
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