Ti-plasmid are widely utilized to introduce DNA into plant cells. but natural form of Ti plasmids are not suitable for as cloning vector because
The size of the Ti plasmids is 150kb-200kb, it is extremely difficult to manipulation.
Binary vector
Consist a helper plasmid and a donor plasmid.
Helper plasmid (binary cloning vector)contains disarmed Ti plasmid, entire T-DNA deletion. vir genes are remained.
Donor plasmid is small E.coli plasmid that carrying a truncated T-DNA region that contains site for excision.
Helper plasmid (binary cloning vector)
- Replication origins for E. coli and Agrobacterium
- A promoter for expression of the target gene
- Selectable markers for bacteria and for plants
- T-DNA border sequence of Ti plasmid
- A cloning site for insertion of the gene of interest
Expression Vectors
- Expression vectors are DNA molecules that can be used to introduce a gene of interest into a host cell and induce its expression. This technology is widely utilized in biotechnology research, allowing researchers to study the function and regulation of genes.
Expression Vectors – components
- A multiple cloning site
- Selectable markers for the identify the transformants
- Replication origin that initiates the replication process
- A promoter for initiation of transcription
- A transcriptional terminator
- A regulatory sequence
- A ribosomal binding site for the initiate translation
- a start codon translation
- A translation terminal site
- Elements for modification – add the sequence for modifying the gene products
