Part:BBa_K5160116
pTRV2-35S promoter - P19-NOS terminator
Overview
To shorten the time required for plant expression validation experiments, we employed the TRV virus for transient infection. By constructing a corresponding vector on the virus, we successfully integrated the target gene into the tomato genome. However, plants possess a gene silencing mechanism as an antiviral defense, which can produce siRNAs to degrade viral RNAs. The P19 protein, as a viral silencing suppressor, can bind to siRNAs, preventing them from binding to and degrading viral RNAs, thereby allowing the virus to replicate and spread smoothly within host cells. Therefore, we incorporated the P19 protein sequence into the TRV2 vector to inhibit the tomato plant's own RNA interference system and ensure that the viral vector could persist in the tomato for a certain period of time. To ensure experimental accuracy, we set up a control group by constructing P19 on the TRV2_35S promoter_P19_NOS terminator.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21INCOMPATIBLE WITH RFC[21]Illegal XhoI site found at 1645
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000INCOMPATIBLE WITH RFC[1000]Illegal BsaI site found at 66
Illegal BsaI site found at 1369
Illegal BsaI.rc site found at 983
Illegal BsaI.rc site found at 1334
Illegal BsaI.rc site found at 1648
CaMV 35S Promoter
The CaMV 35S promoter refers to the 35S promoter derived from the cauliflower mosaic virus (CaMV). This promoter directs the synthesis of 35S RNA during plant infection by CaMV and enables efficient expression in tissues of many dicotyledonous plants. As a constitutive promoter, the CaMV 35S promoter can initiate gene expression in all tissues. It is persistent, with relatively constant RNA and protein expression levels, but lacks spatiotemporal specificity. For detailed information on this component, please refer to BBa_K788000.
P19
The P19 protein, derived from tomato bushy stunt virus (TBSV), can suppress the host's RNA silencing effect on foreign genes, improve the stability of heterologous gene transcripts, and thereby promote the expression of heterologous proteins. It is widely used in transient expression systems for transgenic plants, as well as tobacco, Arabidopsis, and tomato leaves or protoplasts. For detailed information on this component, please refer to BBa_K5160006.
NOS Terminator
The nopaline synthase terminator (NOS terminator) plays an important role in genetic engineering, particularly in plant expression vectors. It ensures that the transcription of the target gene in plants terminates at a specific location, thereby avoiding unnecessary gene expression extension and ensuring the accuracy and efficiency of gene expression. Therefore, it is commonly used to terminate the transcription of target genes in plant vectors, thereby improving expression efficiency. For detailed information on this component, please refer to BBa_P10401.
Source
35S Promoter
The CaMV 35S promoter is derived from the 35S promoter of the cauliflower mosaic virus (CaMV) and is used to initiate the expression of the target gene.
P19
The P19 protein originates from the tomato bushy stunt virus (TBSV) and can suppress the RNA silencing effect of the host on foreign genes, thereby enhancing the stability of heterologous gene transcripts.
NOS Terminator
The nopaline synthase terminator (NOS terminator) primarily originates from Agrobacterium tumefaciens. In genetic engineering, particularly plant genetic engineering, the NOS terminator is often used as an important element in plant expression vectors to mark the end position of target gene transcription.
Design consideration
During the design process, we introduced the silencing suppressor P19 to inhibit the tomato plant's own RNA interference system, ensuring that the viral vector could persist in the tomato for a certain period of time. Considering that the inhibitory effect of the P19 protein is not absolute and may be influenced by various factors such as the concentration of P19, the characteristics of the target gene, and the type of host cells, we deliberately constructed the P19 protein on TRV2_35S promoter_P19_NOS terminator to ensure the accuracy and reliability of the experimental results.
| None |