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Transformation of Intron Hairpin RNAi (ihpRNAi vector) of ERD15 Gene into Tobacco |
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| รหัสดีโอไอ | |
| Creator | Payungsak Rauyaree |
| Title | Transformation of Intron Hairpin RNAi (ihpRNAi vector) of ERD15 Gene into Tobacco |
| Contributor | Aroonothai Sawwa, Paranee Sawangsri, Boonruanrat Rueangwised |
| Publisher | Department of Agriculture |
| Publication Year | 2563 |
| Journal Title | Thai Agricultural Research Journal |
| Journal Vol. | 38 |
| Journal No. | 3 |
| Page no. | 256-266 |
| Keyword | Hairpin RNA, ERD15 gene, gene silencing, ihpRNAi, drought tolerance, intron hairpin RNA vector |
| URL Website | http://at.doa.go.th/journal |
| Website title | Thai Agricultural Research Journal |
| ISSN | 0125-8389 |
| Abstract | ERD15 (Early Responsive to Dehydration 15) is a negative regulator of abscisic acid (ABA) responses which includes rapid activation or response to drought and freeze as well as stomatal closure regulation. It is widely involved in the process of gene transcription and drought tolerance in plants. The objective of this study was to utilize the high-throughput system for making hairpin RNA (hpRNA) constructs of ERD15 gene fragment using one tube restriction-ligation approach. The cloning of the 609 bp full-length synthetic ERD15 gene silencing (GenBank accession number MN816266) flanked with BsaI restriction sites at both sense and antisense using the pRNAi-GG (Golden Gate) based on the Golden Gate (GG) cloning had been successfully constructed and named the synthetic gene pRNAi-ERD15 construct. For Agrobacterium-mediated transformation, the PCR analysis results using 35S CaMV (reverse) and Nos (forward) primers showed that twenty four percent (4 out of 17 calluses) were transformed plants. The ERD15 gene sequence of the transformed tobacco plant when compared with the nucleotide database in GanBank exhibited sequence similarity to that of ERD15 gene. In conclusion, we succeeded in constructing ihpRNAi plasmid construct of the ERD15 gene to be used for plant gene transformation with any other tolerant traits in plants. |
