MCC

 Related Clones for MDH MCC Projects

TbCS  pBMB10 Citrate Synthase from Trypanosoma brucei.  The CS from T brucei is cut into a pGEX4T vector at the XhoI site using Gibson assembly.    The GST is N terminal of the CS and has a thrombin cleavage site with the ss sequence.  The same CS gene has also been cloned without the ss sequence (pBMB12). The expression of either clone has not been tested.  Under construct (DataSheet / SnapGeneFile)

TbcFH pBMB13. Cytosolic Fumarate from T brucei.  Cloned into pGEX4T vector at the XhoI site using Gibson assembly.    The GST is N terminal of fumarase and has a thrombin cleavage site. The expression has not been tested.  Under construct (DataSheet / SnapGeneFile)

TbcFH pBMB14. Mitochondrial Fumarate from T brucei.  Cloned into pGEX4T vector at the XhoI site using Gibson assembly.    The GST is N terminal of fumarase and has a thrombin cleavage site. The expression has not been tested.  Under construct (DataSheet / SnapGeneFile)

hCS  pET28ahCS.  Also called hCS noTEV brain.  Cloned 2014. This is human citrate synthase accession NM-004077.  Initially cloned from human brain cDNA library and cloned into pET28a with NcoI at 5’ and XhoI at 3’.  This will not express in bacterial cultures due to high number of rare codons even if rosetta bacterial strains are used.  Should be avoided and not confused with CS for bacterial expression.  Can be used for subcloning for mammalian expression or other projects. Available upon request

pcDNA3 HA-hCS.  hCS cloned as described above as hCS no TEV brain was cloned into pcDNA2 with an added HA tag at the 5’ end using HindIII and XhoI sites of pcDNA3.1(+).  The final product has an HA-tag at the N-terminal of human citrate synthase and should express in mammalian cells.  The plasmid is 6.8 kbp and is Ampr.  (DataSheet / SnapGeneFile) 

hCS w/TEV  aka CISY_HUMAN (O75390) aka pET28a hCS w/TEV. Human Citrate Synthase.  This clone was codon optimized for expression in bacterial cells and synthesized for PCR cloning into pET28a.  The first 28 amino acids are mitochondrial transit peptide and have not been included in this clone.  (DataSheet / SnapGeneFile)

pGEX4T hCS. The pET28a hCS w/TEV was Gibson assembly cloned into pGET4T.  Expression is not strong. (DataSheet / SnapGeneFile) 

AATC_Human (P17174) aka hGOT1. Human cytosolic (aspartate aminotransferase).  Aka AAT and AST.  Codon optimized and synthesized. Includes amino acids 1-413 with His tag and TEV protease site on N terminus of hGOT1.  Expression or activity has not yet been tested. (DataSheet / SnapGeneFile)

AATM_Human (P00505) aka hGOT2.  Human mitochondrial (aspartate aminotransferase).  Aka AAT and AST.  Codon optimized and synthesized without the first 29 mitochondrial transit residues with an initiator Met and His-TEV on the N terminus. Expression or activity has not yet been tested. (DataSheet / SnapGeneFile)

PCKGC_HUMAN (P35558) aka hPEPCK1 aka PCK1  Human cytosolic phosphoenolpyruvate carboxykinase (GTP).  His tag is C terminal to PEPCK coding. Expression or activity has not yet been tested. (DataSheet / SnapGeneFile)

MAOX_HUMAN (P48163)  aka hME1 Human cytosolic NADP-dependent malic enzyme.  His tag is C terminal to ME1 coding. Expression or activity has not yet been tested. (DataSheet / SnapGeneFile)