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Application of polypeptide in improvement of content of very long chain fatty acids in plants

A fatty acid, ultra-long technology, applied in the field of genetic engineering, to achieve the effect of increasing yield

Active Publication Date: 2019-03-01
INST OF OIL CROPS RES CHINESE ACAD OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Although Zhang Xin et al. (Shandong Agricultural Sciences, 50(6), 2018) cloned the FAE1 gene from the cultivar peanut F112-2, it has not been reported that the gene can increase the content of very long-chain fatty acids in plants. The practical application needs to be further proved

Method used

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  • Application of polypeptide in improvement of content of very long chain fatty acids in plants
  • Application of polypeptide in improvement of content of very long chain fatty acids in plants
  • Application of polypeptide in improvement of content of very long chain fatty acids in plants

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0049] Example 1 Preparation method of peanut fatty acid carbon chain elongation enzyme gene AhyA.KCS1 and AhyB.KCS1

[0050] The developing pods of the peanut variety "Zhonghua 16" were collected, and the developing seeds were taken out and quickly frozen with liquid nitrogen. According to the instructions of the RNA extraction kit (Quanshijin Company, Beijing), the total RNA of peanut developing seeds was extracted. cDNA was obtained according to the operating instructions of the reverse transcription kit (Quanshijin Company, Beijing). Primers were designed according to the gene sequences of Aradu.J6P55 and Araip.S3IU8 in the ancestral diploid wild species Arachis duranensis (AA) and Arachis ipaensis (BB) in the peanut genome database PeanutBase (https: / / peanutbase.org / ). Using the upstream primer 5'-ATGGCTGATGCAAAAGCA-3'(SEQ ID NO:5) and the downstream primer 5'-TCAGATGGCAGATACCCTTGGA-3'(SEQ ID NO:6), using the cDNA of the developing seed as a template, a fragment of about...

Embodiment 2

[0051] Example 2 Structural Analysis of AhyA.KCS1 and AhyB.KCS1 Genes

[0052] Because the cultivar peanut (Arachis hypogaea L.) is allotetraploid (AABB), the AhyA.KCS1 and AhyB.KCS1 genes are the corresponding copies of its A and B genomes, respectively. The DNA coding regions of the AhyA.KCS1 and AhyB.KCS1 genes are both 1533bp, encoding 510 amino acids, and their DNA sequence and amino acid sequence homology all reach more than 99%. Both AhyA.KCS1 and AhyB.KCS1 genes have FAE1_CUT1_RppA and ACP_syn_III_C domains, which are necessary for KCS enzyme function.

Embodiment 3

[0053] Example 3 Production of ultra-long chain saturated fatty acids in yeast cells

[0054] Production of ultra-long chain saturated fatty acids in yeast cells using AhyA.KCS1 or AhyB.KCS1 genes. The AhyA.KCS1 and AhyB.KCS1 genes were respectively linked into the yeast expression vector pYX242, and the pYX242-AhyA.KCS1 and pYX242-AhyB.KCS1 yeast expression vectors were constructed. Such as figure 1 Vector schematic diagram of pYX242-AhyA.KCS1 and pYX242-AhyB.KCS1, TPI is triose phosphate isomerase promoter, Ter is terminator, flori is phage replication origin, LEU2 is LEU2 gene, 2μori is yeast replication origin, Ampicillin is the ampicillin resistance gene, and pUC ori is the replication origin of Escherichia coli.

[0055] pYX242, pYX242-AhyA.KCS1 and pYX242-AhyB.KCS1 were respectively transformed into yeast cells Inv Sci1 (Invitrogen, USA) using a yeast transformation kit (Invitrogen, USA). The transformed yeast was cultured on a medium lacking leucine, and a positive ...

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Abstract

The invention discloses separated peanut beta-ketoacyl-CoA synthase (KCS) genes AhyA.KCS1 (SEQ ID No. 2) and AhyB.KCS1 (SEQ ID No. 4). The invention also provides a method for producing transgenic yeast and plants by using the genes; the method can increase the content of very long chain fatty acids, especially behenic acid, in the plants, and the highest content reaches 12.3%; the total amount ofVLCFA in the yeast is increased to 6.1% from 0.4%, namely, is increased by nearly 15 times; the total amount of the VLCFA in the arabidopsis transgenic plant is increased to 25.0% or above, namely, is increased by 4 times or more. The method has an important application value in the improvement of plant fatty acids and the cultivation and production of high value-added oil crops.

Description

technical field [0001] The invention relates to the field of genetic engineering, in particular to peanut β-ketoacyl-CoA synthase (β-ketoacyl-CoAsynthase, KCS) and its application in plant fatty acid improvement. Background technique [0002] Among the oils consumed by humans, vegetable oil is being paid more and more attention to. Unlike animal oil, which is mainly composed of medium-chain saturated fatty acids, many vegetable oils are rich in long-chain fatty acids. Among them, ultra-long-chain fatty acids are not only essential components of organisms, but also in It has important biological functions in organisms, and has high application value in chemical industry, pharmaceuticals, health care, nutrition, etc., and is an important raw material for production. Using vegetable oil to produce ultra-long-chain fatty acids is an economical, environmentally friendly and sustainable way, but the content of ultra-long-chain saturated fatty acids above 18C in natural vegetable o...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N9/10C12N15/54C12N15/82A01H5/10A01H6/54
CPCC12N9/1029C12N15/8247C12Y203/01041
Inventor 淮东欣廖伯寿薛晓梦雷永晏立英万丽云康彦平
Owner INST OF OIL CROPS RES CHINESE ACAD OF AGRI SCI
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