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Acta Agronomica Sinica ›› 2021, Vol. 47 ›› Issue (6): 1082-1089.doi: 10.3724/SP.J.1006.2021.04116

• SPECIAL SECTION: GENOMICS AND GENETIC IMPROVEMENT IN MAIN BAST FIBER CROPS • Previous Articles     Next Articles

Identification and expression of PAL genes in sisal

HUANG Xing1(), XI Jin-Gen1, CHEN Tao2, QIN Xu2, TAN Shi-Bei1, CHEN He-Long3, YI Ke-Xian1,*()   

  1. 1Environment and Plant Protection Institute, Chinese Academy of Tropical Agricultural Sciences/Key Laboratory of Integrated Pest Management on Tropical Crops, Ministry of Agriculture and Rural Affairs/Hainan Key Laboratory for Monitoring and Control of Tropical Agricultural Pests, Haikou 571101, Hainan, China
    2Guangxi Subtropical Crops Research Institute, Nanning 530001, Guangxi, China
    3Institute of Tropical Bioscience and Biotechnology, Chinese Academy of Tropical Agricultural Sciences, Haikou 571101, Hainan, China
  • Received:2020-05-30 Accepted:2020-09-13 Online:2021-06-12 Published:2020-10-10
  • Contact: YI Ke-Xian E-mail:huangxing@catas.cn;yikexian@126.com
  • Supported by:
    The National Key Research and Development Program of China(2018YFD0201100);The China Agriculture Research System(CARS-16);The Hainan Provincial Natural Science Foundation of China(319QN275);The Hainan Provincial Natural Science Foundation of China(320RC698);The Guangxi Key Research and Development Program(桂科AB18221105);The Belt and Road Tropical Project(BARTP-08)

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Abstract:

Sisal is an important fiber crop in tropical areas, but its research foundation of molecular biology is relatively weak, and the mechanism of fiber development still remains unclear. Phenylalanine ammonia-lyase (PAL) is the first enzyme of lignin bio-synthesis, which is an important component of fiber. According to published transcriptome data, two sisal PAL genes with complete coding sequences were successfully identified. Their expression patterns during sisal leaf development were consistent with previously reported PAL activity changes during fiber development, indicating that PAL was closely related to lignin bio-synthesis. Phylogenetic analysis showed that sisal PALs were closely related with Agave americana. Selection pressure analysis showed similar selection pressure of PALs in sisal and A. americana, which were higher than those in A. tequilana. This might be caused by the convergent evolution of fiber-related traits in sisal and A. americana. In addition, sisal PALs were not significantly expressed under neither copper nor lead stress, which might be caused by post-transcriptional regulation under heavy metal stresses. It was worth noting that the expression of sisal PALs was highly up-regulated after Phytophthora nicotianae Breda inoculation. Sisal PALs might participate in the bio-synthesis of disease resistance-related secondary metabolites in phenylpropanoid pathway, as well as plant cell-wall mediated immunity. Therefore, functional characterization of sisal PALs could improve the understanding of mechanisms in fiber development and disease resistance, which is of great importance for breeding new sisal varieties with high yield, high quality and multiple resistance.

Key words: sisal, PAL gene, phylogenetic analysis, selection pressure, expression pattern, adverse stress

Table 1

Primers of qRT-PCR in the study"

基因
Gene		 正向引物
Forward primer (5°-3°)		 反向引物
Reverse primer (5°-3°)		 产物长度
Product size (bp)
AhPAL1 AGCAGTGATTGGGTGATGGA GAGGAGGGTGTTGATTCGGA 216
AhPAL2 GCGATTGGGAAGCTCATGTT GAGATGAGGCCCAGTGAGTT 239
AhPP2A CCTCCTCCTCCTTCGGTTTG GCCATGAATGTCACCGCAGA 235

Table 2

PAL genes and its protein physicochemical properties and subcellular localization in sisal"

基因
Gene		 碱基长度
Gene length (bp)		 蛋白长度
Protein length (aa)		 蛋白分子量
Molecular weight (kD)		 理论等电点
pI		 亚细胞定位
Subcellular localization
AhPAL1 2112 703 76,474.38 5.85 细胞质(2.462) Cytoplasmic (2.462)
AhPAL2 2118 705 76,267.12 5.93 细胞质(2.245) Cytoplasmic (2.245)

Table 3

Accessions of PALs in Agave"

基因
Gene		 物种
Species		 GenBank序列号
GenBank accession
AhPAL1 Agave H11648 MT536163
AhPAL2 Agave H11648 MT536164
AmPAL1 Agave americana GBHM01016452.1
AmPAL2 Agave americana GBHM01016955.1
AdPAL1 Agave deserti GAHT01019079.1
AdPAL2 Agave deserti GAHT01004501.1
AqPAL1 Agave tequilana GAHU01002518.1
AqPAL2 Agave tequilana GAHU01004755.1

Fig. 1

Phylogenetic analysis of PALs in Arabidopsis, rice, and Agave species At: Arabidopsis thaliana; Ah: Agave H11648; Am: Agave americana; Ad: Agave deserti; Aq: Agave tequilana; Ao: Asparagus officinalis; Lus: Linum usitatissimum; Gorai: Gossypium raimondii; Os: Oryza sativa; GRMZM: Zea mays; Bradi: Brachypodium sylvaticum; Sobic: Sorghum bicolor. Multiple alignment of amino acid sequences was carried out and Neighbor-Joining evolutionary tree was constructed by ClustalX 2.0 software (Bootstrap values were tested for 1000 trails)."

Table 4

Selection pressure analysis of PALs in Agave"

基因
Gene		 物种/物种
Specie/specie		 同义突变频率
Ks		 非同义突变频率
Ka		 非同义突变频率/同义突变频率
Ka/Ks
PAL1 Ad/Ah 0.1176 0.0098 0.083333
Ad/Am 0.2956 0.0155 0.052436
Ad/Aq 0.0179 0.0019 0.106145
PAL2 Ad/Ah 0.3324 0.0271 0.081528
Ad/Am 0.3347 0.0280 0.083657
Ad/Aq 0.0872 0.0129 0.147936

Fig. 2

Selection pressure analysis of PALs in Agave Abbreviations of species are the same as those given in Fig. 1. Sliding window analysis was conducted by DnaSP with a window size of 30 bp and a step size of six bp."

Fig. 3

Transcriptome expression of PALs in Agave Abbreviations of species are the same as those given in Fig. 1."

Fig. 4

Relative expression of AhPALs at different leaf developmental stages and under abotic/biotic stresses in sisal L0: shoot; L1: unexpended leaf; L2: expended leaf; CK: control; CU: copper stress; PB: lead stress; PN: Phytophthora nicotianae Breda inoculation. * and ** indicate significant differences at the 0.05 and 0.01 probability levels, respectively."

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