Scientia Agricultura Sinica ›› 2018, Vol. 51 ›› Issue (21): 4102-4118.doi: 10.3864/j.issn.0578-1752.2018.21.009

• HORTICULTURE • Previous Articles     Next Articles

Identification and Expression Analysis of LBD Gene Family in Grape

HongHong HE(),ZongHuan MA,YuanXia ZHANG,Juan ZHANG,ShiXiong LU,ZhiQiang ZHANG,Xin ZHAO,YuXia WU,Juan MAO()   

  1. College of Horticulture, Gansu Agricultural University, Lanzhou, 730070
  • Received:2018-04-28 Accepted:2018-06-25 Online:2018-11-01 Published:2018-11-01
  • Contact: HongHong HE,Juan MAO E-mail:1152420683@qq.com;maojuan@gsau.edu.cn

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Abstract:

【Objective】The objectives of this research are to identify the Lateral Organ Boundaries Domain(LBD) family genes from grape(Vitis vinifera) genome, to know the profile of LBD family such as gene number, gene structure and abiotic stress expression characteristics in grape, and to provide theoretical basis for exploring what roles the LBD transcription factors play in abiotic stress of grape.【Method】The LBD genes in grape genome-wide were identified by BLAST software in the grape genome database based on LBD genes have been reported from Arabidopsis. DNAMAN5.0, Clustalx, MapInspect, MEME, GSDS2.0, ExPASy and MEGA5.0 software were used for carry out various bioinformatics analysis of LBD. The expression profile chip was draw by the data comes from Affymetrix Gene Chip 16K in PLEXdb. The qRT-PCR was used to detect the expression of grape LBD gene family in abiotic stress.【Result】Total of 30 LBD genes were identified from grape genome, which can be divided into ClassⅠand ClassⅡ. The classⅠcan be divided into 5 subtribe which are namedⅠa,Ⅰb,Ⅰc,Ⅰd andⅠe. ClassⅡ can be divided into Ⅱa and Ⅱb. The number of amino acids is between 127 and 386, and the theoretical equivalence point is between 4.77 and 9.28 in the VvLBD gene family by physicochemical analysis. The analysis of the gene's location on the chromosome revealed that the 30 genes in this family were distributed on 11 of the 19 chromosomes of the grape, and the chromosome 13 contains 7 genes. Multiple sequence alignments and Motif analysis showed that the VvLBD gene family has three conserved domains, namely the sinusoidal structure, the leucine zipper structure, and the Glycine-alanine-serine (GAS) structure. The VvLBD gene family is mainly expressed in chloroplast, mitochondria and nucleus through the sub-cellular location analyzing. The alpha-helix and irregular curl are the main structure of the secondary structure in the VvLBD gene family. The expression of the most gene was increased under the salt and PEG stress, and the expression characteristic was changed by the stress time change in the chip expression profile. The qRT-PCR results showed the expression of VvLBD8, VvLBD11, VvLBD12, VvLBD15, VvLBD16, VvLBD17 were raised to the 3, 1, 8, 4, 5, and 13 times compared the control under the 400 mmol .L -1NaCl treatment. The expression of VvLBD12 and VvLBD19 was raised to the 4 and 26 times compared the control under 10% PEG treatment.【Conclusion】30 LBD gene family members were identified from the grape genome. These members distributed on 11 of the 19 chromosomes of the grape and the evolution of VvLBD is highly conservative. The LBD gene family is related to the abiotic stress, but the different expression were existed between the different members when the encountered the adversity environments.

Key words: Vitis vinifera, LBD gene family, evolutionary analysis, expression profile, qRT-PCR

Table 1

qRT-PCR primers for expression on analysis of VvLBD"

基因Gene 上游引物Forward primer (5′-3′) 下游引物Reverse primer (5′-3′)
VvLBD1 CAGCAGGAGACTGACAGCATGTTC GATAGAAGCCAAGAGTGAGTTCCAACC
VvLBD2 GCCGTGGATGCAGTGCTCAG GCTTGGACTTAGAAGAACGCTTGAATG
VvLBD3 TGGCCTTCATCTCCACCGTACC CCGCTCCGAACACAGGATTCAC
VvLBD4 GAGCTAGTGAACATGCAATGCCAAC GCTGTCATCGAGGAAGAAGGAGTTAC
VvLBD5 AGAGGAGCCTGCCTTACCAACTC AGCTGAGGTAGTGTTGAAGGTTGTTG
VvLBD6 TCCTCCACCTCCTCTACCTCTACC CCTGTTGTGCTTGAAGGCAGTATTG
VvLBD7 GCTGCGGTTACCATTGCTTATGAAG TGTAGATTAACAACCTGCTGCTGGAG
VvLBD8 GCCTTGCAGCAGCAGGTAGC TGCGGCGATACTGAGTTCATGTAAC
VvLBD9 TTCGCTAATGTCCACAAGGTCTTCG ACGGCATCTTCTCGATGTGCAAC
VvLBD10 TTCGTTAATGTCCACAAGGTCTTCGG AATCAGACCAACACAGCCATAGACG
VvLBD11 GTCCTTCATCTCTGCTGTTCCTGAG TCACGGTCCGGCCACACG
VvLBD12 AGCAGCAGCAGCAGCATCAG TCTCTTCCTCGTCTTCGTCCTCATC
VvLBD13 AGTTCTCCATCGTTCACAAGGTCTTC GCAGGTATGATATGGCTCCGACAC
VvLBD14 CAGCACAGCCACCAGCACAG GGAGGAAGAGGAGTTAGGAGGAAGAG
VvLBD15 CTCAACCAACAGGCTCTGCTACTATC AAGGAGGTGGCTGTTCATTCATTAGG
VvLBD16 AGCAGCAGCAGCCACAATATCAG TCCATGACAAGATTGCCATCCTCAAG
VvLBD17 GATGCCTTCTCCTTCTCTTCCAACTG GTTCCTGCAACGCCTTGACAATG
VvLBD18 CTTGCAGCTCTGAGGATGGAATACG AGCTAGAATACAACTCTCATCGCATCG
VvLBD19 ATTCGTGATCCAGTCTATGGTTGTGC ATCTGTCCAGGAAGAAGCTTGCATTAG
VvLBD20 GCCAATGCCACCGTCTTCCTC CACAAGCCTCGTAGAGAAGAGAACTG
VvLBD21 GCCACCAATGCTGACCTGATCC CTCCACCTCCACCTCCATGACTC
VvLBD22 GATGGTAACGCCGCCTTCACTC ACCTCTGGAGCCACCGAACTG
VvLBD23 CCTGTCTATGGCTGTGTTGCTCATG GCGGTGCCAGAGAAGGAATGC
VvLBD24 GTCCGTCAGCGATGGCACATG ATAAGGAATGACCGACGATGAAGAAGG
VvLBD25 CCTTCTTCCAACTTCAACAACTCCTTG GGTCGGATGGAGAGCGGTAGAG
VvLBD26 TGCGGATGAGTTGTAATGGCTGTC GCGAGGAAGACGGTGGCATTG
VvLBD27 TCGTTACGCCTGCAATGAGATGTC GCTTCCTCCTCCTTCATTACCAATCC
VvLBD28 GCAGCGGTGGACACAGTTCTC CAGCGAGTATGCGTCACGTAGC
VvLBD29 TAGGAGAAGATGTGCTCAGGACTGTG TGCTTCATACACCATGCTACTCACTG
VvLBD30 CCAACTCCGCCAACTCCAGATG CTGTGGATGGTGGTGGTGATTGTG
UBI GCTCGCTGTTTTGCAGTTCTAC AACATAGGTGAGGCCGCACTT

Fig. 1

The chromosome location of the LBD gene family in Vitis vinifera"

Table 2

The information of LBD transcription factor family in Vitis vinifera"

基因
Gene		 基因登录号
Gene accession No.		 染色体定位
Chromosome location		 长度
Length
of CDS (bp)		 基因全长
Full length genomic (bp)		 分子量
Molecular
weight (kD)		 等电点
pI		 氨基酸
Amino acid		 外显子
Exon		 cDNA全长
FL-cDNA (bp)		 拟南芥同源基因Arabidopsis homologous gene
登录号
Accession No.		 名称
Gene name		 E值
E-value
VvLBD1 GSVIVT01011895001 Chr.01:3210000..3211373 1158 1374 43.30087 8.96 386 3 1158 AT1G68510 LBD42 7e-05
VvLBD2 GSVIVT01011896001 Chr.01:3202459..3205665 726 1076 26.97486 9.28 242 2 998 AT1G68510 LBD42 7e-05
VvLBD3 GSVIVT01037853001 Chr.03:7098160..7099838 615 1679 22.87014 6.73 205 3 714 AT4G37540 LBD39 1e-09
VvLBD4 GSVIVT01025128001 Chr.06:4180823..4182134 609 1312 22.02887 4.77 202 2 953 AT2G28500 LBD11 8e-14
VvLBD5 GSVIVT01024662001 Chr.06:7971774..7972539 492 766 18.36397 6.40 164 2 644 AT1G31320 LBD4 0.003
VvLBD6 GSVIVT01024592001 Chr.06:8584134..8585865 690 1735 24.92138 8.58 229 3 1529 AT2G30340 LBD13 6e-18
VvLBD7 GSVIVT01028294001 Chr.07:5837981..5838834 699 854 25.72175 5.87 232 2 750 AT2G42430 LBD16 0.018
VvLBD8 GSVIVT01028295001 Chr.07:5846731..5847540 588 810 21.46149 7.54 195 2 588 AT2G42430 LBD16 0.018
VvLBD9 GSVIVT01003547001 Chr.07:14517830..14519654 426 1825 15.69194 8.60 142 2 513 AT5G66870 LBD36 2e-09
VvLBD10 GSVIVT01003548001 Chr.07:14524923..14526703 663 1781 23.96578 9.16 220 2 765 AT5G66870 LBD36 2e-09
VvLBD11 GSVIVT01000141001 Chr.07:15589316..15590604 687 1289 24.96159 8.30 229 2 1167 AT4G37540 LBD39 0.005
VvLBD12 GSVIVT01029979001 Chr.08:2636746..2653598 777 16853 27.99185 5.85 259 4 777 AT3G03760 LBD20 7e-27
VvLBD13 GSVIVT01032752001 Chr.13:1022036..1023024 381 989 14.19636 8.80 127 3 417 AT2G30130 LBD12 4e-09
VvLBD14 GSVIVT01032714001 Chr.13:1310001..1311298 675 1298 24.53103 8.99 225 3 956 AT2G30340 LBD13 2e-11
VvLBD15 GSVIVT01016500001 Chr.13:3355781..3356736 660 955 24.29509 5.08 219 2 849 AT2G42430 LBD16 8e-08
VvLBD16 GSVIVT01016335001 Chr.13:4853462..4854800 594 1339 21.75785 5.67 197 2 677 AT2G28500 LBD11 3e-16
VvLBD17 GSVIVT01016332001 Chr.13:4961089..4962007 498 919 18.35293 8.21 166 2 498 AT2G28500 LBD11 3e-16
VvLBD18 GSVIVT01016329001 Chr.13:4999327..4999957 444 631 16.23288 8.72 148 2 505 AT3G11090 LBD21 8e-04
VvLBD19 GSVIVT01016328001 Chr.13:5001203..5002266 420 1064 15.73712 8.50 140 2 462 AT3G11090 LBD21 8e-04
VvLBD20 GSVIVT01031035001 Chr.14:21210990..21212344 564 1355 20.29311 8.98 187 3 1124 AT1G68510 LBD42 7e-05
VvLBD21 GSVIVT01032415001 Chr.14:27152774..27157752 606 4979 22.07197 8.52 201 3 720 AT3G27650 LBD25 0.003
VvLBD22 GSVIVT01027621001 Chr.15:14983162..14985661 795 2500 28.19431 8.12 264 3 924 AT2G45420 LBD18 2e-21
VvLBD23 GSVIVT01027620001 Chr.15:14992324..14994786 585 2463 21.17114 6.50 195 2 585 AT2G42430 LBD16 1.1
VvLBD24 GSVIVT01018486001 Chr.16:14419741..14420915 672 1175 24.58742 7.65 223 2 672 AT5G66870 LBD36 3e-11
VvLBD25 GSVIVT01010625001 Chr.16:15894465..15895136 462 672 17.36371 9.08 154 2 462 AT3G26620 LBD23 7e-19
VvLBD26 GSVIVT01008284001 Chr.17:3660239..3661508 684 1270 24.77241 8.05 228 3 1035 AT3G02550 LBD41 0.005
VvLBD27 GSVIVT01007677001 Chr.17:10578569..10581053 468 2485 16.93792 6.81 156 3 670 AT5G63090 LOB 3e-22
VvLBD28 GSVIVT01009360001 Chr.18:7746214..7747271 669 1058 24.10543 8.54 222 2 939 AT3G49940 LBD38 3e-07
VvLBD29 GSVIVT01013631001 Chr.Un:1949856..1952420 528 2565 19.30889 7.68 176 2 1646 AT1G16530 LBD3 2e-10
VvLBD30 GSVIVT01006269001 Chr.Un:24370759..24373006 651 2248 23.99094 8.25 217 1 1243 AT1G65620 LBD6 3e-47

Fig. 2

Alignment of conserved domains of LBD family in Vitis vinifera"

Fig. 3

The phylogenetic tree and gene structure of LBD family in Vitis vinifera"

Fig. 4

The unrooted phylogenetic tree of LBD transcription factors in Vitis vinifera(●) and Arabidopsis(○)"

Fig. 5

LBD gene family sequence analysis in Vitis vinifera A: LBD gene family motif analysis in Vitis vinifera; B: LOGO of 8 conserved motif of LOB domain"

Table 3

Subcellular location prediction of LBD gene in Vitis vinifera"

名称
Name		 细胞质
Cytoplasm		 叶绿体
Chloroplast		 细胞核
Nucleus		 线粒体
Mitochondria		 过氧化物
酶体
Peroxisome		 细胞骨架
Cytoskeleton		 液泡
Vacuole		 内质网
Endoplasmic
network		 细胞膜
Plasma membrane		 高尔基体
Golgi apparatus		 细胞基
Extracellular matrix		 细胞核和细胞质
Nuclear and cytoplasmic		 线粒体和细胞质
Mitochondrial and cytoplasmic		 细胞核和细胞膜
Nuclear and plasma membrane
VvLBD1 1.5 0 6.5 4.5 0 0 0 0 0 0 0 4.5 3 0
VvLBD2 2 3 3 6 0 0 0 0 0 0 0 0 0 0
VvLBD3 0 2 8 3 0 0 0 0 0 0 0 0 0 0
VvLBD4 0 5 6 3 0 0 0 0 0 0 0 0 0 0
VvLBD5 1 3 6 1.5 0 0 0 0 1 0 0 0 1.5 0
VvLBD6 6 1 6 0 0 0 0 0 0 0 0 0 0 0
VvLBD7 0 5 8 0 0 0 0 0 0 0 0 0 0 0
VvLBD8 0 8 4 1 0 0 0 0 0 0 0 0 0 0
VvLBD9 0 6 4.5 3 0 0 0 0 0 0 0 3 0 0
VvLBD10 0 7 4 1 0 0 0 0 1 0 0 0 0 0
VvLBD11 0 3 9 2 0 0 0 0 0 0 0 0 0 0
VvLBD12 3 5 2 0 0 2 0 0 1 0 0 0 0 0
VvLBD13 0 12 1 0 0 0 0 0 0 0 0 0 0 0
VvLBD14 0 3 6 4 0 0 0 0 0 0 0 0 0 4.5
VvLBD15 1 2 5 5 0 0 0 0 0 0 0 0 0 0
VvLBD16 1 8 0 4 0 0 0 0 0 0 0 0 0 0
VvLBD17 0 11 2 0 0 0 0 0 0 0 0 0 0 0
VvLBD18 0 4 5.5 4 0 0 0 0 0 0 0 3.5 0 0
VvLBD19 0 13 0 0 0 0 0 0 0 0 0 0 0 0
VvLBD20 0 2 0 11 0 0 0 0 0 0 0 0 0 0
VvLBD21 0 0 13 0 0 0 0 0 0 0 0 0 0 0
VvLBD22 4 3 6 0 0 0 0 0 0 0 0 0 0 0
VvLBD23 0 10 2 1 0 0 0 0 0 0 0 0 0 0
VvLBD24 0 1 4.5 7 0 0 0 0 0 0 0 3 0 0
VvLBD25 0 2 5.5 6 0 0 0 0 0 0 0 3.5 0 0
VvLBD26 0 11 0 2 0 0 0 0 0 0 0 0 0 0
VvLBD27 0 4 8.5 0 0 0 0 0 0 0 0 5 0 0
VvLBD28 0 5 6.5 1 0 0 0 0 0 0 0 4 0 0
VvLBD29 0 11 2 0 0 0 0 0 0 0 0 0 0 0
VvLBD30 0 3 9 2 0 0 0 0 0 0 0 0 0 0

Table 4

The secondary structure of LBD protein sequence in Vitis vinifera"

名称Name α-螺旋Alpha helix (%) β-转角Beta turn (%) 不规则卷曲Random coil (%)
VvLBD1 39.38 6.74 36.01
VvLBD2 20.66 6.20 53.72
VvLBD3 37.56 8.78 39.02
VvLBD4 45.05 2.97 41.58
VvLBD5 41.46 1.83 39.63
VvLBD6 49.34 3.49 37.12
VvLBD7 48.28 6.90 38.79
VvLBD8 39.49 7.69 38.97
VvLBD9 33.80 8.45 30.28
VvLBD10 56.36 4.55 27.37
VvLBD11 32.89 11.84 40.35
VvLBD12 33.98 3.47 44.02
VvLBD13 52.76 0.00 26.77
VvLBD14 43.11 4.44 44.89
VvLBD15 47.95 6.85 33.33
VvLBD16 29.95 5.58 41.62
VvLBD17 39.76 2.41 42.77
VvLBD18 37.84 6.76 41.22
VvLBD19 45.71 6.43 37.14
VvLBD20 27.81 11.23 38.50
VvLBD21 31.34 9.95 42.79
VvLBD22 33.71 6.82 49.24
VvLBD23 51.79 7.69 33.33
VvLBD24 39.91 8.97 32.29
VvLBD25 51.30 6.49 28.57
VvLBD26 32.46 10.96 38.60
VvLBD27 37.82 7.69 45.51
VvLBD28 41.11 12.96 33.33
VvLBD29 42.61 4.55 42.05
VvLBD30 49.77 5.99 36.41

Fig. 6

Expression profile of LBD gene in Vitis vinifera Dark blue, light blue, colorless, light red and dark red are used to represent gene expression levels. Blue indicates weak gene expression, red indicates strong gene expression"

Fig.7

VvLBD gene expression under different treatment A: Control; B: 24 h treatment by 400 mmol·L-1 NaCl; C: 24 h treatment by 50 μmol·L-1 ABA; D: 24 h treatment by 10%PEG"

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