Biotechnology Bulletin ›› 2024, Vol. 40 ›› Issue (2): 160-171.doi: 10.13560/j.cnki.biotech.bull.1985.2023-0856
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GONG Li-li1,2(), YU Hua2, YANG Jie2, CHEN Tian-chi2, ZHAO Shuang-ying2, WU Yue-yan1,2()
Received:
2023-09-04
Online:
2024-02-26
Published:
2024-03-13
Contact:
WU Yue-yan
E-mail:gongllstudy@163.com;wyy2000@zwu.edu.cn
GONG Li-li, YU Hua, YANG Jie, CHEN Tian-chi, ZHAO Shuang-ying, WU Yue-yan. Identification and Analysis of Grape(Vitis vinifera L.)CYP707A Gene Family and Functional Verification to Fruit Ripening[J]. Biotechnology Bulletin, 2024, 40(2): 160-171.
引物名称 Primer name | 引物序列 Primer sequence(5'-3') |
---|---|
Vitvi07g01751-F | ATGCTGCTGAAGAAGCCACA |
Vitvi07g01751-R | TCAGCATTCCTTCCAAAACTTAGC |
35S:VvCYP707A-F | ACGAGCTCGGTACCCGGGATGCTGCTGAAGAAGCCACAG |
35S:VvCYP707A-R | GGGCGAATTGGTCGACGCATTCCTTCCAAAACTTAGCAGG |
Table 1 Primer information
引物名称 Primer name | 引物序列 Primer sequence(5'-3') |
---|---|
Vitvi07g01751-F | ATGCTGCTGAAGAAGCCACA |
Vitvi07g01751-R | TCAGCATTCCTTCCAAAACTTAGC |
35S:VvCYP707A-F | ACGAGCTCGGTACCCGGGATGCTGCTGAAGAAGCCACAG |
35S:VvCYP707A-R | GGGCGAATTGGTCGACGCATTCCTTCCAAAACTTAGCAGG |
基因Gene | 正向引物 Forward primer(5'-3') | 反向引物 Reverse primer(5'-3') |
---|---|---|
Vitvi02g01269 | F: GTGTTCTGCAAGCTGTCACG | R: TGGCATCTTCCCAAGTCAGAG |
Vitvi03g00508 | F: ATCGGCCTTTGACATGGGAT | R: AGTGGCATCACCTTCCAACC |
Vitvi06g00498 | F: GCAGCTTCTCAACGAGACTTTCAGG | R: GAATTCCCAGCAATCCACCTCCTTG |
Vitvi07g01751 | F: CTCTGCGGCTCTATTCCCAG | R: CTTGGTGGGTATGTGGGCTT |
Vitvi18g00792 | F: TTGTCCTGGAAATGAGCTTGCCAAG | R: CAATTCCACCTTGTGATCCCACCAC |
Actin | F: CAAGAGAAACCATCCCTAGCTG | R: TCAATCTGTCTAGGAAAGGAAG |
VvPYL1 | F: CAAGAGAAACCATCCCTAGCTG | R: TCAATCTGTCTAGGAAAGGAAG |
VvPYL2 | F: TCCCTACTGTCTGGTCCGTC | R: CCAATCTCTCCGTGCTGGTC |
VvPYL3 | F: GACGAGTACATCCGCAGACA | R: TGATGTGCTTGACGAGAGAGG |
VvPYL4 | F: GTGCTCTTCCCTACTCGCTC | R: GAAGCGATGAACGACGGAAC |
VvPYL5 | F: ATAGACGGCTGTGATGGGTG | R: TGAACTGCACTGGTTCTCCC |
VvPYL6 | F: CCACTACCAGCACTGAAAGGT | R: TCGTCCTTGGTGTTTCCCTC |
Table 2 Primers for RT-qPCR
基因Gene | 正向引物 Forward primer(5'-3') | 反向引物 Reverse primer(5'-3') |
---|---|---|
Vitvi02g01269 | F: GTGTTCTGCAAGCTGTCACG | R: TGGCATCTTCCCAAGTCAGAG |
Vitvi03g00508 | F: ATCGGCCTTTGACATGGGAT | R: AGTGGCATCACCTTCCAACC |
Vitvi06g00498 | F: GCAGCTTCTCAACGAGACTTTCAGG | R: GAATTCCCAGCAATCCACCTCCTTG |
Vitvi07g01751 | F: CTCTGCGGCTCTATTCCCAG | R: CTTGGTGGGTATGTGGGCTT |
Vitvi18g00792 | F: TTGTCCTGGAAATGAGCTTGCCAAG | R: CAATTCCACCTTGTGATCCCACCAC |
Actin | F: CAAGAGAAACCATCCCTAGCTG | R: TCAATCTGTCTAGGAAAGGAAG |
VvPYL1 | F: CAAGAGAAACCATCCCTAGCTG | R: TCAATCTGTCTAGGAAAGGAAG |
VvPYL2 | F: TCCCTACTGTCTGGTCCGTC | R: CCAATCTCTCCGTGCTGGTC |
VvPYL3 | F: GACGAGTACATCCGCAGACA | R: TGATGTGCTTGACGAGAGAGG |
VvPYL4 | F: GTGCTCTTCCCTACTCGCTC | R: GAAGCGATGAACGACGGAAC |
VvPYL5 | F: ATAGACGGCTGTGATGGGTG | R: TGAACTGCACTGGTTCTCCC |
VvPYL6 | F: CCACTACCAGCACTGAAAGGT | R: TCGTCCTTGGTGTTTCCCTC |
基因ID Gene ID | 氨基酸长度 Amino acid length/aa | 分子量大小 Molecular weight/kD | 等电点 pI | 亚细胞定位 Subcellular location |
---|---|---|---|---|
Vitvi02g01269 | 470 | 53 118.2 | 9.03 | 内质网 |
Vitvi03g00508 | 479 | 54 538.9 | 9.68 | 内质网 |
Vitvi06g00498 | 488 | 55 496.0 | 10.00 | 内质网 |
Vitvi07g01751 | 447 | 50 689.3 | 9.50 | 内质网 |
Vitvi18g00792 | 470 | 53 448.7 | 10.08 | 内质网 |
Table 3 Information of CYP707A proteins in grape
基因ID Gene ID | 氨基酸长度 Amino acid length/aa | 分子量大小 Molecular weight/kD | 等电点 pI | 亚细胞定位 Subcellular location |
---|---|---|---|---|
Vitvi02g01269 | 470 | 53 118.2 | 9.03 | 内质网 |
Vitvi03g00508 | 479 | 54 538.9 | 9.68 | 内质网 |
Vitvi06g00498 | 488 | 55 496.0 | 10.00 | 内质网 |
Vitvi07g01751 | 447 | 50 689.3 | 9.50 | 内质网 |
Vitvi18g00792 | 470 | 53 448.7 | 10.08 | 内质网 |
Fig. 2 Phylogenetic tree analysis of CYP707A protein The phylogenetic tree was constructed with the full-length amino acid sequences of the 24 CYP707A using MEGA X. The analysis included 5 Vitis vinifera VvCYP707A proteins, 2 Solanum lycopersicum SlCYP707A proteins, 4 Arabidopsis thaliana AtCYP707A proteins, 2 Daucus carota DcCYP707A proteins, 4 Oryza sativa OsCYP707A proteins, 2 Malus domestica MdCYP707A proteins, 2 Solanum tuberosum StCYP707A proteins and 4 Citrus CsCYP707A proteins. The VvCYP707A proteins in V. vinifera were marked with red color and red stars
Fig. 5 Physiological parameters and VvCYP707As related expression related to fruit development in two kinds of V. vinifera A: ‘Yin Hong’ and ‘Yong Zaohong’ at different developmental periods; S1-S2: young fruit stage; S3: expansion stage; S4: veraison stage; S5: mature stage. The same below. B-D: ‘Yin Hong’ and ‘Yong Zaohong’ fruit single weight, fruit hardness and soluble sugar content. The data used are the mean ±SD of the three biological replicates. * on the column indicates significant differences between the two varieties, *: P<0.05, **: P<0.01, ***: P<0.001. E: VvCYP707A gene expression changes in ‘Yin Hong’ and ‘Yong Zaohong’. Different lowercase letters represent a significant difference(P<0.05); n=3, the same below
Fig. 6 VvCYP707As expression patterns in different tissues and different periods in ‘Yin Hong’ A: Flesh; B: pericarp; C: tissue specificity of VvCYP707As expression in veraison stage
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