Cloning and Functional Analysis of Potato Light-trapping Pigment Binding Protein Gene StCP24 under Different Light Treatments

doi:10.7668/hbnxb.20195132

Abstract

Abstract:

In order to study the role of phototrapping pigment binding protein gene in potato photosynthesis,the phototrapping pigment binding protein gene StCP24 (LOC102586836)was bioinformatics analyzed and cloned into potato variety Dongnong 310 to obtain transgenic potato.The morphological indexes,leaf physiological indexes,fluorescence parameters,and gene relative expression of transgenic and control lines were measured under different light intensities after clonal propagation.The function of StCP24 gene was analyzed.The effect of this gene on chloroplasts was determined by ultrastructural observation.The results of bioinformatics analysis showed that StCP24 protein was a hydrophilic protein,and the promoter of StCP24 gene contained elements such as photoresponse,defense and stress response.Phylogenetic analysis showed that the amino acids encoded by StCP24 gene in potato were closely related to those encoded by StCP24 gene in cultivated tomato.Under different light intensity treatments,StCP24 gene overexpressed potato significantly increased stem diameter,leaf area,leaf weight,root length,chlorophyll a,b content,Fv/Fm,ETR,ETRmax and qP compared with wild-type potato.The overexpression of StCP24 gene can make the lamellar stacking of thylakoid granules more tightly and to a higher degree.In conclusion,the overexpression of StCP24 gene can increase the content of chlorophyll a and b in leaves,improve the electron transfer rate in photo cooperative photoreaction,and promote the growth of transgenic potato plants.

Key words: Potato, StCP24 gene, Light intensity, Overexpression

LIU Yulin, LI Shiwei, PEI Yating, GAO Hongxiu, TANG Xinhua, SHI Ying. Cloning and Functional Analysis of Potato Light-trapping Pigment Binding Protein Gene StCP24 under Different Light Treatments[J]. Acta Agriculturae Boreali-Sinica, 2024, 39(6): 55-63. doi: 10.7668/hbnxb.20195132.

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Figures/Tables 13

Tab.1 Primer information

引物名称 Primer name	引物序列(5'—3') Primer sequence (5'—3')	用途 Purpose
StCP24-F	CGGGATCCATGGCTACAACATCTGCTGCGGT	基因CDS扩增
StCP24-R	GCTCTAGACAAACCAAGAGCTCCCAGGGGGG
35S+StCP24-F	AATTCGGTCCCCAGATTAGCCT	转基因马铃薯验证
35S+StCP24-R	CAAACCAAGAGCTCCCAGGGGG
StCP241-F	TTGAAATGGTACAGAGAGG	qRT-PCR
StCP241-R	AGAAGTGTGCCAAAAGAGA
Elfα-F	GATGTTGTGCCAAAGGATGT	内参基因
Elfα-R	AACTTGGTCAATGCGAGA

Tab.2 Prediction of secondary structure of StCP24 protein

二级结构 Secondary structure	氨基酸数目/个 Amino acid number
α-螺旋Alpha helix	99
链的延伸Extended strand	23
β-转角Beta turn	6
无规卷曲Random coil	128

Fig.1 Prediction of secondary structure(A)and tertiary structure(B)of StCP24 protein

Fig.2 StCP24 promoter element ARE.Anaerobic induction;circadian.Regulate circadian rhythm control;TCA-element.Salicylic acid response;TGA-element.Auxin response;TC-rich repeats.Defense and stress response;ABRE.Abscisic acid response;GATA-motif,Sp1,TCCC-motif,chs-CMA2a,G-box.Photoresponse;LTR.Low temperature response.

Fig.3 Phylogenetic tree of the amino acid of potato StCP24

Fig.4 Electrophoresis of plasmid PCR and double enzyme digestion A:M.Marker,1.Empty carrier,2—3.Recombinant plasmid;B:1.Plasmid,2.Double enzyme digested plasmid.

Fig.5 Electrophoretic image of PCR detection of transgenic potatoes M.Marker;1.Positive control;2.Negative control;3.ddH₂O;4—7.PCR product of 35S+CP24 primer amplification.

Fig.6 The growth comparison between StCP24 overexpression strain and WT

Fig.7 Phenotypic analysis of transgenic and non-transgenic potatoes under different light conditions

Fig.8 Chlorophyll analysis of transgenic and non-transgenic potatoes under different light conditions

Fig.9 Fluorescence parameter analysis of transgenic and non-transgenic potatoes under different light conditions

Fig.10 The expression heat map of StCP24 under different light treatments

Fig.11 Chloroplast ultrastructure of StCP24 overexpression and control potato C.Chloroplast;G.Granular lamella.

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