What is A TOPO cloning vector?
What is A TOPO cloning vector?
TOPO cloning is a molecular biology technique in which DNA fragments are cloned into specific vectors without the requirement for DNA ligases. Taq polymerase has a nontemplate-dependent terminal transferase activity that adds a single deoxyadenosine (A) to the 3′-end of the PCR products.
What is TOPO PCR cloning?
Topoisomerase based cloning (TOPO cloning) is a DNA cloning method that does not use restriction enzymes or ligase, and requires no post-PCR procedures. Sounds easy right? The technique relies on the basic ability of complementary basepairs adenine (A) and thymine (T) to hybridize and form hydrogen bonds.
Is TOPO cloning technique directional what is directional and how is this achieved?
Directional TOPO cloning enables cloning of blunt-ended PCR products in a 5´→3´ orientation directly into a expression vector using a 5-minute ligation reaction, thereby eliminating subcloning steps and saving you time. Topoisomerase I then ligates the PCR product in the correct orientation. …
Is TOPO cloning technique directional?
In practise TOPO-cloning is not directional, so the insert is equally likely to ligate in both directions. However, in Geneious only the current orientation (defined by the orientations of documents in the sequence viewer) will be ligated into the vector.
What is directional TOPO cloning?
Directional TOPO cloning enables cloning of blunt-ended PCR products in a 5´→3´ orientation directly into a expression vector using a 5-minute ligation reaction, thereby eliminating subcloning steps and saving you time. Topoisomerase I then ligates the PCR product in the correct orientation.
Is Topo cloning technique directional?
What enzyme is specifically associated with the TOPO cloning kit and how does it work?
The key to TOPO cloning is the enzyme DNA topoisomerase I, which functions both as a restriction enzyme and as a ligase. Its biological role is to cleave and rejoin DNA during replication.
What are the features of Zero® Blunt® Topo® PCR cloning kits?
Features of the Zero® Blunt® TOPO® PCR Cloning Kits: Simple — no ligase, post-PCR procedures, or PCR primers containing specific sequences are required. Competent cells: Various options—choose from kits with either general, high-efficiency, bacteriophage T1-resistant, fast-growing competent cells, or use your own.
Can TOPO TA and Zero Blunt vectors be used for in vitro transcription?
Yes, the TOPO TA cloning and Zero Blunt cloning vectors can be used for in vitro transcription. The T7, SP6, and T3 promoters contained in these vectors are active phage promoters, and the corresponding polymerases will transcribe RNA from these promoters.
What is the PCR™Blunt II-TOPO® vector used for?
The pCR™Blunt II-TOPO® vector is designed to clone blunt-ended PCR products generated by thermostable proofreading polymerases such as Platinum® Pfx DNA Polymerase. The pCR™Blunt II-TOPO® vector contains: • EcoRI sites flanking the PCR product insertion site for easy excision of inserts
What is ZeroZero blunt® Topo®?
Zero® Blunt® TOPO® kits are available with a variety of competent cells, or without competent cells, depending on your needs and budget. Features of the Zero® Blunt® TOPO® PCR Cloning Kits: Simple — no ligase, post-PCR procedures, or PCR primers containing specific sequences are required.
