Looking for breakthrough ideas for innovation challenges? Try Patsnap Eureka!

Persimmon proanthocyanidin precursor transmembrane transport gene dkmate5 and its application

A technology of transmembrane transport and proanthocyanidins, applied in application, genetic engineering, plant gene improvement, etc., can solve the problems of less research on transmembrane transport, differences in astringent types of persimmon varieties, and incomplete clarity

Active Publication Date: 2022-02-11
HUAZHONG AGRI UNIV
View PDF4 Cites 0 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, the structural genes and regulatory genes of proanthocyanidin biosynthesis have been cloned and identified, but the transmembrane transport of proanthocyanidin precursors is less studied
In particular, whether the preferential transport of persimmon proanthocyanidin precursors causes the differences in the type of astringency of persimmon varieties is not yet fully clear.

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • Persimmon proanthocyanidin precursor transmembrane transport gene dkmate5 and its application
  • Persimmon proanthocyanidin precursor transmembrane transport gene dkmate5 and its application
  • Persimmon proanthocyanidin precursor transmembrane transport gene dkmate5 and its application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0019] Example 1 Acquisition and Analysis of DkMATE5 and DkMATE4 Gene Sequences

[0020] 1. Acquisition of DkMATE5 and DkMATE4 gene sequences in the fruit of 'E Shi No. 1'

[0021] (1) First, download the HMM information of the MATE gene family from the Pfam database (http: / / pfam.xfam.org), and use the HMMER software to compare the persimmon genome (http: / / www.kakiwi.zju.edu.cn / cgi- bin / persimmon / index.cgi) to obtain candidate MATE protein sequences, compare the candidate protein sequences to the Pfam database, and retain the MATE genes with conserved domains as DkMATE family gene members, use the transcriptome database to search for DkMATE differentially expressed genes, and determine which Differential expression of DkMATE4 and DkMATE5 genes;

[0022] (2) Utilize the Primer Premier 5.0 software to design RACE primers therebetween, wherein the RACE primers used to amplify the DkMATE4 gene are shown in SEQ ID NO.13-14, and the RACE primers used to amplify the DkMATE5 gene are...

Embodiment 2

[0034] Example 2 Analysis of transient transformation of DkMATE4 and DkMATE5 in persimmon leaves

[0035] (1) According to the DkMATE5 gene sequence shown in SEQ ID NO.1 obtained in Example 1 and the DkMATE4 gene sequence shown in SEQ ID NO.12, respectively design the MATE5-OEF shown in SEQ ID NO.8-9 and MATE5-OER and MATE4-OEF and MATE4-OER as shown in SEQ ID NO.19-20, and according to the instructions, use the Gateway method to introduce the DkMATE5 gene and DkMATE4 gene into the entry vector pDONR207, and then introduce the overexpression through the LR reaction In the vector pK2GW7, overexpression recombinant plasmids were respectively constructed, and the overexpression recombinant plasmids were transformed into Agrobacterium (GV3101) by conventional methods.

[0036] (2) The above-mentioned transfected Agrobacterium was instantly transformed into the leaves of 'Eshi 1' by conventional injection infiltration method, and the leaves infected by Agrobacterium were collected ...

Embodiment 3

[0038] Example 3 Escherichia coli functional complementation experiment to determine the transport preference of DkMATE5

[0039] 1. Construction of PGEX-6P-1-DkMATE5 expression vector

[0040] (1) Use PrimeSTAR Max Premix (2x) high-fidelity enzyme (TaKaRa, Japan), use the full-length plasmid of the DkMATE5 gene as a template, and use primers containing BamH I and Not I restriction sites (such as SEQ ID NO.10 -11), amplified and cloned into the prokaryotic expression vector PGEX-6P-1 to construct the prokaryotic expression plasmid PGEX-6P-1-DkMATE5.

[0041] 2. Escherichia coli functional complementation experiment

[0042] The prokaryotic expression plasmid PGEX-6P-1-DkMATE5 constructed above was transformed into Escherichia coli mutant strain acrB lacking drug efflux ability according to conventional methods. Pick a successfully transformed single colony into 5mL LB liquid medium, shake at 200r / min at 37°C for about 12h. Take 500 μL of bacterial liquid into 50 mL of LB li...

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

PUM

No PUM Login to View More

Abstract

The invention discloses a persimmon proanthocyanidin precursor transmembrane transport gene DkMATE5 and its application, and belongs to the technical field of plant genetic engineering. The invention obtains DkMATE4 and DkMATE5 genes through the first screening of a transcriptome database, and finds that the DkMATE5 protein has a proanthocyanidin transport function. The analysis of in vivo complementation experiments in Escherichia coli further proved that the DkMATE5 gene promotes the preferential transmembrane transport of persimmon proanthocyanidin precursors catechin, epicatechin and epicatechin gallate. The invention provides a scientific basis and new genetic resources for the analysis of the differences in the composition and content of proanthocyanidins in different types of persimmons that have lost their astringency and for the genetic improvement of sweet persimmons.

Description

technical field [0001] The invention belongs to the technical field of plant genetic engineering, and specifically relates to persimmon proanthocyanidin precursor transmembrane transport gene DkMATE5 and application thereof. Background technique [0002] Persimmon fruit contains a large amount of Proanthocyanidins (PAs; commonly known as "persimmon tannin"), which is an important indicator of fruit maturity and commodity quality. The biosynthesis of proanthocyanidins involves shikimic acid, public phenylpropanoid, core flavonoid-anthocyanidins and proanthocyanidin-specific pathways. Proanthocyanidin precursors are synthesized in the cytoplasm, recognized by transport proteins located on the tonoplast membrane and transported to the vacuole for polymerization, thereby forming macromolecular proanthocyanidins. At present, the structural genes and regulatory genes of proanthocyanidin biosynthesis have been cloned and identified, but the transmembrane transport of proanthocyani...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

Application Information

Patent Timeline
no application Login to View More
Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/29C12N15/82C12N15/11C12Q1/6851A01H5/00A01H6/00
CPCC07K14/415C12Q1/6851C12N15/8243C12Q2531/113C12Q2563/107
Inventor 徐莉清刘颖罗正荣张青林陈文兴郭大勇
Owner HUAZHONG AGRI UNIV
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Patsnap Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Patsnap Eureka Blog
Learn More
PatSnap group products

Browse by: Latest US Patents, China's latest patents, Technical Efficacy Thesaurus, Application Domain, Technology Topic, Popular Technical Reports.

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap|About US| Contact US: help@patsnap.com