Biotechnology Bulletin ›› 2021, Vol. 37 ›› Issue (9): 106-113.doi: 10.13560/j.cnki.biotech.bull.1985.2021-0287
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HU Zi-yao(), DAI Pei-hong, LIU Chao, Madina Mulati, WANG Qian, Wugalihan Abuduwili, ZHAO Yi, SUN Ling, XU Shi-jia, LI Yue()
Received:
2021-03-09
Online:
2021-09-26
Published:
2021-10-25
Contact:
LI Yue
E-mail:1916938224@qq.com;liyue6905@126.com
HU Zi-yao, DAI Pei-hong, LIU Chao, Madina Mulati, WANG Qian, Wugalihan Abuduwili, ZHAO Yi, SUN Ling, XU Shi-jia, LI Yue. Molecular Cloning,Expression and VIGS Construction of a Small GTP-binding Protein Gene GhROP3 in Gossypium hirsutum[J]. Biotechnology Bulletin, 2021, 37(9): 106-113.
基因 Gene | 序列 Sequence(5'-3') | 用途 Application |
---|---|---|
GhROP3 | F-ATGAGCACTGCAAGATTTATC R-TCAAAGGAAAGCACATGTTCT | 基因克隆 |
qGhROP3 | F-GGATCCCAGAGCTAAGACAT R-CTCCCTGAGATGTTGATATTGGTG | qRT-PCR |
qGhUBQ7 | F-GAAGGCATTCCACCTGACCAAC R-CTTGACCTTCTTCTTCTTGTGCTTG | qRT-PCR |
VIGS-GhROP3 | F-TCTAGAGGATCCCAGAGCTAAGACATTA R-CCCGGGTTGCAAGGCTTTCTCTTTGG | VIGS载 体构建 |
VIGS-qGhCLA1 | F-GCCCTTTGTGCATCTTC R-CTCTAGGGGCATTGAAG | qRT-PCR |
VIGS-qGhROP3 | F-TTCTCCTTTTGCAAGGCTTTCTC R-TGACAGACCATCCTAACGCA | qRT-PCR |
Table 1 Primer and application
基因 Gene | 序列 Sequence(5'-3') | 用途 Application |
---|---|---|
GhROP3 | F-ATGAGCACTGCAAGATTTATC R-TCAAAGGAAAGCACATGTTCT | 基因克隆 |
qGhROP3 | F-GGATCCCAGAGCTAAGACAT R-CTCCCTGAGATGTTGATATTGGTG | qRT-PCR |
qGhUBQ7 | F-GAAGGCATTCCACCTGACCAAC R-CTTGACCTTCTTCTTCTTGTGCTTG | qRT-PCR |
VIGS-GhROP3 | F-TCTAGAGGATCCCAGAGCTAAGACATTA R-CCCGGGTTGCAAGGCTTTCTCTTTGG | VIGS载 体构建 |
VIGS-qGhCLA1 | F-GCCCTTTGTGCATCTTC R-CTCTAGGGGCATTGAAG | qRT-PCR |
VIGS-qGhROP3 | F-TTCTCCTTTTGCAAGGCTTTCTC R-TGACAGACCATCCTAACGCA | qRT-PCR |
基因 Gene | 开放阅读框 ORF/bp | 氨基酸 Amino acid | 相对分子质量 Relative molecular quality/kD | 理论等电点 Theoretical isoelectric point | 平均疏水性 Average hydrophobicity | 信号肽 Signal peptide | 亚细胞定位 Subcellular localization | 脂肪系数 Fat coefficient |
---|---|---|---|---|---|---|---|---|
GhROP3 | 591 | 196 | 21.75 | 9.45 | -0.138 | 无None | 细胞膜Cytomembrane | 86.02 |
Table 2 Bioinformatics analysis of GhROP3 in cotton
基因 Gene | 开放阅读框 ORF/bp | 氨基酸 Amino acid | 相对分子质量 Relative molecular quality/kD | 理论等电点 Theoretical isoelectric point | 平均疏水性 Average hydrophobicity | 信号肽 Signal peptide | 亚细胞定位 Subcellular localization | 脂肪系数 Fat coefficient |
---|---|---|---|---|---|---|---|---|
GhROP3 | 591 | 196 | 21.75 | 9.45 | -0.138 | 无None | 细胞膜Cytomembrane | 86.02 |
Fig. 2 Expression pattern analysis of GhROP3 A:Salt stress. B:Drought stress. C:Cold stress. D:Verticillium wilt. E:Expression of GhROP3 in different tissues of cotton. The difference between the lowercase letters showed significant difference in different sample time at the 0.05 probability level
Fig. 3 Construction of VIGS vector of GhROP3 A:PCR amplification of target fragment of GhROP3;B:identification of TRV﹕GhROP3 VIGS vectors by restriction enzyme digestion. M:Trans 2K Plus Ⅱ DNA marker
Fig. 4 Verification of VIGS system A:The phenotype of silenced GhCLA1 in cotton;B:silencing efficiency of GhCLA1; C:silencing efficiency of GhROP3. **:Significant difference at the 0.01 probability
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