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Acta Agronomica Sinica ›› 2024, Vol. 50 ›› Issue (10): 2515-2527.doi: 10.3724/SP.J.1006.2024.34149

• CROP GENETICS & BREEDING·GERMPLASM RESOURCES·MOLECULAR GENETICS • Previous Articles     Next Articles

Divergent evolutionary pattern of P5CS gene family and drought tolerance verification of LusP5CS1 in linseed

WANG Ling1(), ZHANG Yan-Ping2, QI Yan-Ni3, WANG Lei1, LI Yu-Xiao1, TAN Mei-Lian1,*(), WANG Wei1,*()   

  1. 1Oil Crops Research Institute, Chinese Academy of Agricultural Sciences / Key Laboratory of Biology and Genetic Improvement of Oil Crops, Ministry of Agriculture and Rural Affairs, Wuhan 430062, Hubei, China
    2Institute of Biotechnology, Gansu Academy of Agricultural Science, Lanzhou 730070, Gansu, China
    3Institute of Crop, Gansu Academy of Agricultural Science, Lanzhou 730070, Gansu, China
  • Received:2023-09-05 Accepted:2024-06-20 Online:2024-10-12 Published:2024-07-08
  • Contact: *E-mail: wangwei03@caas.cn;E-mail: meiliantan@126.com
  • Supported by:
    China Agriculture Research System of MOF and MARA(CARS-14-1-17);Science and Technology Innovation Project of the Chinese Academy of Agricultural Sciences(CAAS- ASTIP-2016-OCRI);National Infrastructure for Crop Germplasm Resources(NCGRC-2023-016)

Abstract:

Pyrroline-5-carboxylate synthetase (P5CS), a crucial enzyme encoded by the P5CS gene, plays a key role in the drought stress response in plants, primarily by regulating proline biosynthesis. In this study, we used two P5CS genes from Arabidopsis thaliana as query sequences to meticulously screen and identify members of the P5CS gene family from the genomes of various important grain and oil crops, including linseed, rapeseed, soybean, peanut, sunflower, rice, and wheat. By thoroughly analyzing the selection pressure, specific sites, and functional differentiation potential of P5CS genes across these diverse crops, we gained a comprehensive understanding of their evolutionary patterns. Furthermore, we validated the functions of these genes in Arabidopsis thaliana, a model organism. Our findings revealed significant differences in gene structure and evolutionary patterns among members of the linseed P5CS gene family compared to their homologs in other crops. Notably, overexpressing the positively selected linseed LusP5CS1 gene in Arabidopsis thaliana resulted in a substantial increase in proline accumulation and enhanced drought resistance in the transgenic plants. Interestingly, even under non-drought stress conditions, the transgenic Arabidopsis thaliana exhibited a notable fitness advantage. This groundbreaking study not only enhances our understanding of the molecular mechanisms underlying drought resistance but also provides a solid theoretical foundation for breeding drought-tolerant linseed varieties.

Key words: linseed, P5CS gene family, phylogenetic analysis, gene duplication, functional divergence

Table 1

Primers used in this study"

引物名称
Primer name
引物序列
Primer sequence (5'-3')
用途
Purpose
LusP5CS1-F ATGGAAGATCCAGTTGGTC 基因克隆
Gene cloning
LusP5CS1-R CTATACAGTTATATCCTTGTGGGTG
LusP5CS1-qF CGCGGAGGGCCTTGTCTTA 荧光定量
qRT-PCR
LusP5CS1-qR GATCGCCTGGCCTCTTTT
pRI201-AN-LusP5CS1-F tcttcactgttgata CATATGATGGAAGATCCAGTTGGTCATGTA 过表达载体构建
Construction of overexpression vector
pRI201-AN-LusP5CS1-R ttgctaatcgacggg CATATGCTATACAGTTATATCCTTGTGGGTGTAGA
AtActin-F TGGCCGATGGTGAGGATATT 拟南芥内参基因
Reference gene of Arabidopsis
AtActin-R AACGGCCTGAATGGCAACAT

Table 2

Members of the P5CS gene family in different plant species"

物种 Species 基因ID Gene ID
拟南芥Arabidopsis AT2G39800 (AtP5CS1), AT3G55610 (AtP5CS2)
胡麻Linseed Lus10001016 (LusP5CS3), Lus10001018 (LusP5CS1), Lus10004696 (LusP5CS7), Lus10004697 (LusP5CS8), Lus10030158 (LusP5CS4), Lus10040262 (LusP5CS2), Lus10040263 (LusP5CS5), Lus10040264 (LusP5CS6)
油菜Rapeseed Brara.C02032 (BnP5CS3), Brara.D00386 (BnP5CS5), Brara.D02414 (BnP5CS4), Brara.E00599 (BnP5CS1), Brara.I03811 (BnP5CS2)
大豆Soybean Glyma.01G099800 (GmP5CS1), Glyma.02G251100 (GmP5CS3), Glyma.03G069400 (GmP5CS5), Glyma.07G137300 (GmP5CS2), Glyma.14G065600 (GmP5CS4), Glyma.18G034300 (GmP5CS6), Glyma.18G188000 (GmP5CS7)
花生Peanut arahy.Tifrunner.gnm1.ann1.WS4P7I (AhP5CS3), arahy.Tifrunner.gnm1.ann1.1W477Y (AhP5CS6), arahy.Tifrunner.gnm1.ann1.AXAR8E (AhP5CS1), arahy.Tifrunner.gnm1.ann1.NA5QFN (AhP5CS4), arahy.Tifrunner.gnm1.ann1.SCHW0S (AhP5CS5), arahy.Tifrunner.gnm1.ann1.V8868B (AhP5CS2)
向日葵Sunflower HanXRQChr07g0201741 (HaP5CS1), HanXRQChr08g0221191 (HaP5CS2)
水稻Rice Os01g62900 (OsP5CS1), Os05g38150 (OsP5CS2)
小麦Wheat Traes_3DL_3E215D878 (TaP5CS1), Traes_3B_C4683D0FA (TaP5CS3), Traes_1DL_0BB66CF71 (TaP5CS4), Traes_1AL_49393CDA7 (TaP5CS5), Traes_3AL_302AF461E (TaP5CS6), Traes_1BL_31105367B (TaP5CS2)

Table S1

Homologous relationships among members of the P5CS gene family in different plant species"

亚家族
Subfamily
基因
Gene
直系同源基因
Orthologous genes
P5CS1 AtP5CS1 AhP5CS3, AhP5CS1, AhP5CS4, BnP5CS3, BnP5CS4, BnP5CS1, GmP5CS5, GmP5CS1, GmP5CS3, GmP5CS4, GmP5CS6, OsP5CS1, TaP5CS1, TaP5CS6, TaP5CS3, LusP5CS6, LusP5CS7, LusP5CS2, HaP5CS1
AhP5CS3 GmP5CS1, GmP5CS6, GmP5CS3, BnP5CS1, OsP5CS1, TaP5CS3, TaP5CS6, HaP5CS1, LusP5CS2, LusP5CS6
AhP5CS5
AhP5CS1
AhP5CS4 GmP5CS6, GmP5CS3, HaP5CS1, BnP5CS1, OsP5CS1, TaP5CS3, TaP5CS6, LusP5CS2, LusP5CS6
BnP5CS3
BnP5CS4 OsP5CS1, GmP5CS1, GmP5CS6, GmP5CS3, TaP5CS3, TaP5CS6, HaP5CS1, LusP5CS2, LusP5CS6
BnP5CS1
GmP5CS5 AhP5CS3, BnP5CS1, OsP5CS1, TaP5CS3, TaP5CS6, HaP5CS1, LusP5CS2, LusP5CS6
GmP5CS1
GmP5CS3
GmP5CS4 HaP5CS1, BnP5CS1, OsP5CS1, TaP5CS3, TaP5CS6, LusP5CS2, LusP5CS6
GmP5CS6 GmP5CS1, GmP5CS6, GmP5CS3, BnP5CS1, TaP5CS3, TaP5CS6, HaP5CS1, LusP5CS2, LusP5CS6
OsP5CS1
TaP5CS1
TaP5CS6 OsP5CS1, GmP5CS1, GmP5CS6, GmP5CS3, HaP5CS1, LusP5CS2, LusP5CS6
TaP5CS3 GmP5CS1, GmP5CS6, GmP5CS3, BnP5CS1, OsP5CS1, HaP5CS1
LusP5CS6
LusP5CS7 GmP5CS1, GmP5CS6, GmP5CS3, BnP5CS1, OsP5CS1, TaP5CS3, TaP5CS6, HaP5CS1
LusP5CS2
HaP5CS1
P5CS2 AtP5CS2 AhP5CS2, AhP5CS6, BnP5CS5, BnP5CS2, GmP5CS2, GmP5CS7, OsP5CS2, TaP5CS5, TaP5CS4, TaP5CS2, HaP5CS2, LusP5CS5, LusP5CS8, LusP5CS3, LusP5CS4, LusP5CS1
AhP5CS2 GmP5CS2, GmP5CS7, OsP5CS2, TaP5CS2, BnP5CS2, HaP5CS2, LusP5CS4, LusP5CS1, LusP5CS5
AhP5CS6
BnP5CS5
BnP5CS2 GmP5CS2, GmP5CS7, OsP5CS2, TaP5CS2, HaP5CS2, LusP5CS4, LusP5CS1, LusP5CS5
GmP5CS2 OsP5CS2, TaP5CS2, HaP5CS2, LusP5CS4, LusP5CS1, LusP5CS5
GmP5CS7
OsP5CS2 TaP5CS2, HaP5CS2, LusP5CS4, LusP5CS1, LusP5CS5
TaP5CS5
TaP5CS4
TaP5CS2 HaP5CS2, LusP5CS4, LusP5CS1, LusP5CS5
HaP5CS2 LusP5CS4, LusP5CS1, LusP5CS5
LusP5CS5
LusP5CS8
LusP5CS3
LusP5CS4
LusP5CS1

Fig. 1

Conserved motifs and functional domains of P5CS"

Fig. S1

Phylogenetic relationships of P5CS using Maximum Likelihood method"

Fig. 2

Phylogenetic relationships of P5CSs in different plants Phylogenetic tree constructed using protein sequences of P5CS genes. a: an unrooted tree; b: a rooted proportional polar tree. Leguminous plants (soybean, peanut) are denoted by blue blocks, cruciferous plants (Arabidopsis thaliana, canola) by purple blocks, linseed plants (linseed) by rose red blocks, compositae (sunflower) by orange blocks, and monocotyledonous grasses plants (rice and wheat) by yellow blocks."

Fig. 3

Whole genome and P5CS gene duplication in linseed Linseed whole-genome replication analysis using linseed genome data. a: gene duplication among the 15 chromosomes of linseed with each point representing a replicated gene, red points indicate best homology while blue points denote partial homology, grayed-out points represent other genes; b: linseed whole genome duplication; c: duplication within the linseed P5CS gene family."

Table S2

Replication analysis of P5CS genes"

物种树节点
Species tree node
基因树节点
Gene tree node
支持率
Support
类型
Type
基因1
Genes 1
基因2
Genes 2
N1 n1 0.125 Hannuus_HanXRQChr12g0369501 Hannuus_HanXRQChr08g0221191 Lusitatissimum_Lus10001016
Lusitatissimum_Lus10005468
Lusitatissimum_Lus10001019
Lusitatissimum_Lus10030158
Lusitatissimum_Lus10001018
Lusitatissimum_Lus10040262
Lusitatissimum_Lus10004696
Brapa_Brara.D02414.1.p
Brapa_Brara.C02032.1.p
Brapa_Brara.E00599.1.p
Athaliana_AT2G39800.1
Athaliana_AT3G55610.1
Brapa_Brara.D00386.1.p
Brapa_Brara.I03811.1.p
Hannuus_HanXRQChr06g0177501
Gmax_Glyma.07G137300.1.p
Gmax_Glyma.18G188000.1.p
Taestivum_Traes_1DL_2E85568A1.2
Ahypogaea_arahy.Tifrunner.gnm1.ann1.V8868B.1
Ahypogaea_arahy.Tifrunner.gnm1.ann1.1W477Y.1
Ahypogaea_arahy.Tifrunner.gnm1.ann1.02CEIS.1
Osativa_LOC_Os01g62900.1
Taestivum_Traes_3AL_30C0103A0.1
Taestivum_Traes_3B_C4683D0FA.2
Taestivum_Traes_3DL_3E215D878.2
Taestivum_Traes_3AL_302AF461E.1
Osativa_LOC_Os05g38150.2
Taestivum_Traes_1AL_49393CDA7.1
Taestivum_Traes_1DL_0BB66CF71.1
Taestivum_Traes_1BL_31105367B.1
Taestivum_Traes_1AL_014F29DA6.1
Ahypogaea_arahy.Tifrunner.gnm1.ann1.WS4P7I.1
Ahypogaea_arahy.Tifrunner.gnm1.ann1.AXAR8E.1
Gmax_Glyma.01G099800.10.p
Gmax_Glyma.03G069400.2.p
Taestivum_Traes_7DS_9330C9BFD.1
Lusitatissimum_Lus10040263
Lusitatissimum_Lus10004697
Lusitatissimum_Lus10040264
Ahypogaea_arahy.Tifrunner.gnm1.ann1.NA5QFN.1
Ahypogaea_arahy.Tifrunner.gnm1.ann1.SCHW0S.1
Gmax_Glyma.02G251100.1.p
Gmax_Glyma.14G065600.1.p
Gmax_Glyma.18G034300.1.p
Hannuus_HanXRQChr07g0201741
Hannuus n2 1 Terminal Hannuus_HanXRQChr12g0369501 Hannuus_HanXRQChr08g0221191
N1 n3 0.625 Lusitatissimum_Lus10001016 Lusitatissimum_Lus10005468 Lusitatissimum_Lus10001019
Lusitatissimum_Lus10030158
Lusitatissimum_Lus10001018
Lusitatissimum_Lus10040262
Lusitatissimum_Lus10004696
Brapa_Brara.D02414.1.p
Brapa_Brara.C02032.1.p
Brapa_Brara.E00599.1.p
Athaliana_AT2G39800.1
Athaliana_AT3G55610.1
Brapa_Brara.D00386.1.p
Brapa_Brara.I03811.1.p
Hannuus_HanXRQChr06g0177501
Gmax_Glyma.07G137300.1.p
Gmax_Glyma.18G188000.1.p
Taestivum_Traes_1DL_2E85568A1.2
Ahypogaea_arahy.Tifrunner.gnm1.ann1.V8868B.1
Ahypogaea_arahy.Tifrunner.gnm1.ann1.1W477Y.1
Ahypogaea_arahy.Tifrunner.gnm1.ann1.02CEIS.1
Osativa_LOC_Os01g62900.1
Taestivum_Traes_3AL_30C0103A0.1
Taestivum_Traes_3B_C4683D0FA.2
Taestivum_Traes_3DL_3E215D878.2
Taestivum_Traes_3AL_302AF461E.1
Osativa_LOC_Os05g38150.2
Taestivum_Traes_1AL_49393CDA7.1
Taestivum_Traes_1DL_0BB66CF71.1
Taestivum_Traes_1BL_31105367B.1
Taestivum_Traes_1AL_014F29DA6.1
Ahypogaea_arahy.Tifrunner.gnm1.ann1.WS4P7I.1
Ahypogaea_arahy.Tifrunner.gnm1.ann1.AXAR8E.1
Gmax_Glyma.01G099800.10.p
Gmax_Glyma.03G069400.2.p
Taestivum_Traes_7DS_9330C9BFD.1
Lusitatissimum_Lus10040263
Lusitatissimum_Lus10004697
Lusitatissimum_Lus10040264
Ahypogaea_arahy.Tifrunner.gnm1.ann1.NA5QFN.1
Ahypogaea_arahy.Tifrunner.gnm1.ann1.SCHW0S.1
Gmax_Glyma.02G251100.1.p
Gmax_Glyma.14G065600.1.p
Gmax_Glyma.18G034300.1.p
Hannuus_HanXRQChr07g0201741
Lusitatissimum n5 1 Terminal Lusitatissimum_Lus10001016 Lusitatissimum_Lus10005468
Lusitatissimum_Lus10001019
Lusitatissimum_Lus10030158
Lusitatissimum_Lus10001018
Lusitatissimum_Lus10040262
Lusitatissimum_Lus10004696
Lusitatissimum n6 1 Terminal Lusitatissimum_Lus10001016 Lusitatissimum_Lus10005468
Lusitatissimum_Lus10001019
Lusitatissimum_Lus10030158
Lusitatissimum_Lus10001018
Lusitatissimum n7 1 Terminal Lusitatissimum_Lus10001016 Lusitatissimum_Lus10005468
Lusitatissimum_Lus10001019
Lusitatissimum_Lus10030158
Lusitatissimum n8 1 Terminal Lusitatissimum_Lus10005468 Lusitatissimum_Lus10001019
Lusitatissimum_Lus10030158
Lusitatissimum n9 1 Terminal Lusitatissimum_Lus10001019 Lusitatissimum_Lus10030158
Lusitatissimum n10 1 Terminal Lusitatissimum_Lus10040262 Lusitatissimum_Lus10004696
N7 n12 1 STRIDE Brapa_Brara.D02414.1.p Brapa_Brara.C02032.1.p
Brapa_Brara.E00599.1.p
Athaliana_AT2G39800.1
Athaliana_AT3G55610.1
Brapa_Brara.D00386.1.p
Brapa_Brara.I03811.1.p
Brapa n14 1 Terminal Brapa_Brara.D02414.1.p, Brapa_Brara.C02032.1.p
Brapa_Brara.E00599.1.p
Brapa n15 1 Terminal Brapa_Brara.D02414.1.p Brapa_Brara.C02032.1.p
Brapa n17 1 Terminal Brapa_Brara.D00386.1.p Brapa_Brara.I03811.1.p
N1 n18 0.25 Hannuus_HanXRQChr06g0177501 Gmax_Glyma.07G137300.1.p Gmax_Glyma.18G188000.1.p
Taestivum_Traes_1DL_2E85568A1.2
Ahypogaea_arahy.Tifrunner.gnm1.ann1.V8868B.1
Ahypogaea_arahy.Tifrunner.gnm1.ann1.1W477Y.1
Ahypogaea_arahy.Tifrunner.gnm1.ann1.02CEIS.1
Osativa_LOC_Os01g62900.1
Taestivum_Traes_3AL_30C0103A0.1
Taestivum_Traes_3B_C4683D0FA.2
Taestivum_Traes_3DL_3E215D878.2
Taestivum_Traes_3AL_302AF461E.1
Osativa_LOC_Os05g38150.2
Taestivum_Traes_1AL_49393CDA7.1
Taestivum_Traes_1DL_0BB66CF71.1
Taestivum_Traes_1BL_31105367B.1
Taestivum_Traes_1AL_014F29DA6.1
Ahypogaea_arahy.Tifrunner.gnm1.ann1.WS4P7I.1
Ahypogaea_arahy.Tifrunner.gnm1.ann1.AXAR8E.1
Gmax_Glyma.01G099800.10.p
Gmax_Glyma.03G069400.2.p
N1 n20 0.125 Gmax_Glyma.07G137300.1.p Gmax_Glyma.18G188000.1.p Taestivum_Traes_1DL_2E85568A1.2
Ahypogaea_arahy.Tifrunner.gnm1.ann1.V8868B.1
Ahypogaea_arahy.Tifrunner.gnm1.ann1.1W477Y.1
Ahypogaea_arahy.Tifrunner.gnm1.ann1.02CEIS.1
Osativa_LOC_Os01g62900.1
Taestivum_Traes_3AL_30C0103A0.1
Taestivum_Traes_3B_C4683D0FA.2
Taestivum_Traes_3DL_3E215D878.2
Taestivum_Traes_3AL_302AF461E.1
Osativa_LOC_Os05g38150.2
Taestivum_Traes_1AL_49393CDA7.1
Taestivum_Traes_1DL_0BB66CF71.1
Taestivum_Traes_1BL_31105367B.1
Taestivum_Traes_1AL_014F29DA6.1
Gmax n22 1 Terminal Gmax_Glyma.07G137300.1.p Gmax_Glyma.18G188000.1.p
Ahypogaea n24 1 Terminal Ahypogaea_arahy.Tifrunner.gnm1.ann1.V8868B.1 Ahypogaea_arahy.Tifrunner.gnm1.ann1.1W477Y.1
Ahypogaea_arahy.Tifrunner.gnm1.ann1.02CEIS.1
Ahypogaea n25 1 Terminal Ahypogaea_arahy.Tifrunner.gnm1.ann1.V8868B.1 Ahypogaea_arahy.Tifrunner.gnm1.ann1.1W477Y.1
N3 n26 1 STRIDE Osativa_LOC_Os01g62900.1 Taestivum_Traes_3AL_30C0103A0.1
Taestivum_Traes_3B_C4683D0FA.2
Taestivum_Traes_3DL_3E215D878.2
Taestivum_Traes_3AL_302AF461E.1
Osativa_LOC_Os05g38150.2
Taestivum_Traes_1AL_49393CDA7.1
Taestivum_Traes_1DL_0BB66CF71.1
Taestivum_Traes_1BL_31105367B.1
Taestivum_Traes_1AL_014F29DA6.1
Taestivum n28 1 Terminal Taestivum_Traes_3AL_30C0103A0.1 Taestivum_Traes_3B_C4683D0FA.2
Taestivum_Traes_3DL_3E215D878.2
Taestivum_Traes_3AL_302AF461E.1
Taestivum n29 1 Terminal Taestivum_Traes_3AL_30C0103A0.1 Taestivum_Traes_3B_C4683D0FA.2
Taestivum_Traes_3DL_3E215D878.2
Taestivum n30 1 Terminal Taestivum_Traes_3B_C4683D0FA.2 Taestivum_Traes_3DL_3E215D878.2
Taestivum n32 1 Terminal Taestivum_Traes_1AL_49393CDA7.1 Taestivum_Traes_1DL_0BB66CF71.1
Taestivum_Traes_1BL_31105367B.1
Taestivum_Traes_1AL_014F29DA6.1
Taestivum n33 1 Terminal Taestivum_Traes_1DL_0BB66CF71.1 Taestivum_Traes_1BL_31105367B.1
Taestivum_Traes_1AL_014F29DA6.1
Taestivum n34 1 Terminal Taestivum_Traes_1BL_31105367B.1 Taestivum_Traes_1AL_014F29DA6.1
Ahypogaea n36 1 Terminal Ahypogaea_arahy.Tifrunner.gnm1.ann1.WS4P7I.1 Ahypogaea_arahy.Tifrunner.gnm1.ann1.AXAR8E.1
Gmax n37 1 Terminal Gmax_Glyma.01G099800.10.p Gmax_Glyma.03G069400.2.p
Lusitatissimum n41 1 Terminal Lusitatissimum_Lus10040263 Lusitatissimum_Lus10004697
Lusitatissimum_Lus10040264
Lusitatissimum n42 1 Terminal Lusitatissimum_Lus10040263 Lusitatissimum_Lus10004697
Ahypogaea n44 1 Terminal Ahypogaea_arahy.Tifrunner.gnm1.ann1.NA5QFN.1 Ahypogaea_arahy.Tifrunner.gnm1.ann1.SCHW0S.1
Gmax n45 1 Terminal Gmax_Glyma.02G251100.1.p Gmax_Glyma.14G065600.1.p
Gmax_Glyma.18G034300.1.p

Table 3

Selection sites analysis of P5CS gene"

物种
Species
基因亚家族 Gene subfamilies
P5CS1 P5CS2
拟南芥 Arabidopsis N/A N/A
胡麻 Linseed 356, 357, 361, 365, 366, 367, 368, 369, 379, 382, 383, 394, 396, 397, 401, 402, 405, 406, 410, 414, 415, 418, 421, 422, 425, 459, 462, 540, 551, 555, 581, 600, 604, 607, 608, 609, 611, 613, 614, 629, 630, 634, 637, 650, 652, 655, 659, 660, 661, 664, 666, 667, 669, 672, 884, 885, 889, 892, 896, 900, 902, 904, 905, 906, 907 125, 383, 396, 405, 422, 424, 555, 611, 614, 650, 669, 884, 889, 892, 904, 906
油菜 Rapeseed 584 620
大豆 Soybean 35, 38, 49 635
花生 Peanut 770,775 408
向日葵 Sunflower 455 N/A
水稻 Rice N/A N/A
小麦 Wheat 126 181, 184

Fig. 4

Selection pressure analysis of the P5CS gene family"

Table 4

Functional differentiation analysis of P5CS gene in linseed"

亚家族1
Subfamily 1
亚家族2
Subfamily 2
θ
θ-value
亚家族1
Subfamily 1
亚家族2
Subfamily 2
θ
θ-value
P5CS1 (胡麻)
P5CS1 (Linseed)
P5CS1 (大豆)
P5CS1 (Soybean)
0.32±0.05 P5CS2 (胡麻)
P5CS2 (Linseed)
P5CS1 (大豆)
P5CS1 (Soybean)
0.30±0.04
P5CS1 (油菜)
P5CS1 (Rapeseed)
0.24±0.05 P5CS1 (油菜)
P5CS1 (Rapeseed)
0.26±0.08
P5CS1 (花生)
P5CS1 (Peanut)
0.36±0.09 P5CS1 (花生)
P5CS1 (Peanut)
0.25±0.07
P5CS1 (向日葵)
P5CS1 (Sunflower)
0.35±0.04 P5CS1 (向日葵)
P5CS1 (Sunflower)
0.27±0.09
P5CS1 (拟南芥)
P5CS1 (Arabidopsis)
0.29±0.04 P5CS1 (拟南芥)
P5CS1 (Arabidopsis)
0.24±0.05
P5CS1 (水稻)
P5CS1 (Rice)
0.46±0.08 P5CS1 (水稻)
P5CS1 (Rice)
0.49±0.06
P5CS1 (小麦)
P5CS1 (Wheat)
0.49±0.05 P5CS1 (小麦)
P5CS1 (Wheat)
0.32±0.08
P5CS2 (大豆)
P5CS2 (Soybean)
0.42±0.05 P5CS2 (大豆)
P5CS2 (Soybean)
0.28±0.05
P5CS2 (油菜)
P5CS2 (Rapeseed)
0.29±0.07 P5CS2 (油菜)
P5CS2 (Rapeseed)
0.43±0.09
P5CS2 (花生)
P5CS2 (Peanut)
0.17±0.09 P5CS2 (花生)
P5CS2 (Peanut)
0.39±0.05
P5CS2 (向日葵)
P5CS2 (Sunflower)
0.25±0.08 P5CS2 (向日葵)
P5CS2 (Sunflower)
0.25±0.04
P5CS2 (拟南芥)
P5CS2 (Arabidopsis)
0.21±0.04 P5CS2 (拟南芥)
P5CS2 (Arabidopsis)
0.37±0.04
P5CS2 (水稻)
P5CS2 (Rice)
0.18±0.07 P5CS2 (水稻)
P5CS2 (Rice)
0.40±0.02
P5CS2 (小麦)
P5CS2 (Wheat)
0.19±0.06 P5CS2 (小麦)
P5CS2 (Wheat)
0.48±0.08
P5CS2 (胡麻)
P5CS2 (Linseed)
0.25±0.07

Table S3

Functional differentiation analysis of P5CS gene in different plant species"

亚家族
Subfamily
物种
Species
θ值θ value
大豆
Soybean
油菜
Rapeseed
向日葵
Sunflower
拟南芥
Arabidopsis
水稻
Rice
小麦
Wheat
P5CS1 花生 Peanut 0.04 ± 0.0.7 0.07 ± 0.05 0.06 ± 0.10 0.03 ± 0.03 0.20 ± 0.05 0.24 ± 0.11
大豆 Soybean 0.03 ± 0.07 0.05 ± 0.07 0.06 ± 0.04 0.16 ± 0.08 0.27 ± 0.08
油菜 Rapeseed 0.04 ± 0.07 0.02 ± 0.04 0.11 ± 0.04 0.28 ± 0.15
向日葵 Sunflower 0.03 ± 0.04 0.20 ± 0.08 0.18 ± 0.07
拟南芥 Arabidopsis 0.24 ± 0.08 0.24 ± 0.11
水稻 Rice 0.03 ± 0.07
P5CS2 花生 Peanut 0.02 ± 0.11 0.05 ± 0.07 0.05 ± 0.08 0.08 ± 0.07 0.21 ± 0.15 0.22 ± 0.15
大豆 Soybean 0.06 ± 0.07 0.07 ± 0.11 0.07 ± 0.05 0.26 ± 0.11 0.22 ± 0.08
油菜 Rapeseed 0.08 ± 0.07 0.07 ± 0.06 0.25 ± 0.14 0.18 ± 0.11
向日葵 Sunflower 0.06 ± 0.04 0.22 ± 0.08 0.20 ± 0.14
拟南芥 Arabidopsis 0.21 ± 0.06 0.18 ± 0.07
水稻 Rice 0.05 ± 0.07

Fig. S2

Detection of LusP5CS1 gene expression in Arabidopsis plants a: Detection of LusP5CS1 gene expression in Arabidopsis plants by RT-PCR; b: Detection of LusP5CS1 gene expression in Arabidopsis plants by qRT-PCR. M: 2000 bp DNA Marker; NC: Negative control. WT: Wild type Arabidopsis; LusP5CS1-OE: Overexpressed LusP5CS1 gene Arabidopsis. The red curve represents transgenic Arabidopsis, and the green curve represents the wild-type control. The inserted bar chart represents the expression level of the LusP5CS1 gene (n=6)."

Fig. 5

Phenotypic changes of Arabidopsis thaliana with overexpression of LusP5CS1 gene a-c: the phenotypic differences between overexpressed LusP5CS1 gene and wild type Arabidopsis under non-drought stress (a, b) and drought stress (c). d-f: the statistical results of proline content (d), biomass (e), and seed weight (f) under drought and non-drought stress, respectively. WT: wild type Arabidopsis; LusP5CS1-OE: overexpressed LusP5CS1 gene Arabidopsis. All data points are means ± standard error (n = 5); *: P < 0.05; **: P < 0.01."

Table S4

Enhanced phenotypic analysis of transgenic Arabidopsis thaliana subjected to drought and non-drought stress conditions"

表型
Phenotype
非干旱胁迫 Non-drought stress 干旱胁迫 Drought stress
野生型
WT
过表达LusP5CS1基因
LusP5CS1-OE
野生型
WT
过表达LusP5CS1基因
LusP5CS1-OE
脯氨酸含量 Proline content (μg g-1) 19.06 ± 4.37 58.30 ± 16.43 48.94 ± 19.05 206.48 ± 60.05
种子重量 Seed weight (g) 0.14 ± 0.02 0.23 ± 0.03 0.08 ± 0.02 0.15 ± 0.02
生物量鲜重 Fresh weight (g) 1.83 ± 0.36 3.00 ± 0.35 1.08 ± 0.11 2.04 ± 0.10
生物量干重 Dry weight (g) 0.19 ± 0.04 0.35 ± 0.08 0.14 ± 0.02 0.34 ± 0.10
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