pTrcHis-TOPO (linearized)
Linearized bacterial vector with 3'-T overhangs and bound topoisomerase, for TOPO® TA cloning and inducible expression of N-terminally 6xHis-tagged proteins.
Sequence Author: Thermo Fisher (Invitrogen)
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Efficient cleavage requires at least two copies of the PluTI recognition sequence. |
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| * Blocked by Dcm methylation. Efficient cleavage requires at least two copies of the NarI recognition sequence. |
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EcoRV is reportedly more prone than its isoschizomer Eco32I to delete a base after cleavage. |
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ApaI can be used between 25°C and 37°C. |
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Sticky ends from different BanII sites may not be compatible. |
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Sticky ends from different BstEII sites may not be compatible.BstEII is typically used at 60°C, but is 50% active at 37°C. |
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| * Blocked by Dam methylation. BclI is typically used at 50-55°C, but is 50% active at 37°C. |
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Sticky ends from different PflMI sites may not be compatible. |
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Sticky ends from different PfoI sites may not be compatible. |
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The 1-base overhangs produced by PflFI may be hard to ligate.Sticky ends from different PflFI sites may not be compatible. |
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The 1-base overhangs produced by Tth111I may be hard to ligate.Sticky ends from different Tth111I sites may not be compatible. |
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Efficient cleavage with AccI requires ≥13 bp on each side of the recognition sequence.Sticky ends from different AccI sites may not be compatible. |
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Prolonged incubation with NdeI may lead to removal of additional nucleotides. |
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Sticky ends from different BspQI sites may not be compatible. |
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Sticky ends from different SapI sites may not be compatible.SapI gradually settles in solution, so a tube of SapI should be mixed before removing an aliquot. |
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PciI is inhibited by nonionic detergents. |
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Sticky ends from different AlwNI sites may not be compatible. |
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Efficient cleavage requires at least two copies of the BfuAI recognition sequence. Sticky ends from different BfuAI sites may not be compatible.BfuAI is typically used at 50°C, but is 50% active at 37°C. |
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Efficient cleavage requires at least two copies of the BspMI recognition sequence. Sticky ends from different BspMI sites may not be compatible. |
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Sticky ends from different BtgI sites may not be compatible. |
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Sticky ends from different StyI sites may not be compatible. |
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| * Blocked by Dam methylation. |
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After cleavage, BamHI-HF® (but not the original BamHI) can remain bound to DNA and alter its electrophoretic mobility. |
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