Difference between revisions of "User:Etienne Robillard/Notebook/Agent Ecoli:Plasmid cloning vectors for gene therapy with phosphohistidine based oligos"

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(Orthogonal DNA synthesis with Benzoyl-derived phosphotriester protecting groups (dA))
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= Plasmid cloning vectors for gene therapy with phosphohistidine-based oligos =
== Eukariotic DNA Promoters ==
* N-6-benzoyl-deoxyadenosine phosphoramidite - Notice the P-N bound indicating the linkage of a phosphonate diester to a benzoyl atom :)
* N-3-oxo-hexanoyl-L-homoserine lactone - A light-inducible phosphohistidine-bound AHL ? A(denosine)TP/cAMP dependent.
* Reminder:
** NHC = (N heterocycle carbene) - Chemical compounds with a N chiral prodrug switch - highly enantioselective molecules
** N = indicates a Nitrogen atom
** P = a Phosphonate (inorganic state) - a compound for oligo synthesis using the [http://en.wikipedia.org/wiki/Oligonucleotide_synthesis#Nucleoside_phosphoramidites nucleoside phosphoramidite] method. 
** P-N = phosphoramidate (a Nitrogen atom bound to a Phosphonate ester in nucleophilic substitution).
deoxynucleotide synthesis:
* cDNA synthesis building block = deoxynucleoside + 3'-phosphonate = synthetic nucleotide analog (dATP) [http://dwb4.unl.edu/Teacher/NSF/C08/C08Links/www.dur.ac.uk/~dbl0www/Staff/Croy/cDNAfigs.htm 1]
oligodeoxyribonucleotide synthesis:
* short 50-200(?) bp restriction endonucleases enzymes for cDNA catalysis/plasmid vector cloning in ''E. coli''
* building blocks for oligonucleotides assembly with phosphoramidites method [http://en.wikipedia.org/wiki/Oligonucleotide 1]
* see also [http://openwetware.org/wiki/CH391L/S12/GeneandGenomeSynthesis#Oligonucleotide_Synthesis Oligonucleotide Synthesis]
* 1 mutation, 1 oligo assay (phosphoramidite-based oligo synthesis): [http://openwetware.org/wiki/Reviews/Directed_evolution/Library_construction#Directing_diversity:_Oligonucleotide-based_methods]
[http://openwetware.org/wiki/20.109(S07):Start-up_genome_engineering#Part_3:_Digest_M13K07 M13K07] cloning in ''E. coli'':
* amino-acid sequence signature (Myc epitope tag) = EQKLISEEDL
=== Orthogonal DNA synthesis with Benzoyl-derived phosphotriester protecting groups (dA) ===
* note! note! Important: Can you elucidate the reasoning of using an carboxilic acid as protecting group for DNA assembly?
*** Bz = Benzoyl (Bz, benzoic acid ester) : 2nd step in the catalysis of Benzilic acid from oxidation of Benzoic acid. See "benzoic acid". A very common [reagent] for site-directed oligonucleotide synthesis.
*** Note: Consensus promoter sequences are by definition strong promoters!
*** 2-cyanoethyl : for protecting phosphoramidites based nucleosides?  XXX
*** araBAD XXX
*** phospho'''tri'''ester = a '''tri'''cyclic N-derived nitrogenous base bound to a Histidine residue made for '''DNA''' synthesis.
* '''Thus phosphohistidine based phosphoramidates can be classified as purine analogues; the substance (Purines) was formally discovered by Emil Fischer during WW1 as a fundamental group of heterocyclic amino acids.'''
** see also [http://en.wikipedia.org/wiki/Piperidine Piperidine] : "Piperidine is also commonly used in chemical degradation reactions, such as the sequencing of DNA in the cleavage of particular modified nucleotides. Piperidine is also commonly used as a base for the deprotection of Fmoc-amino acids used in '''solid-phase peptide synthesis.'''"
== References ==
* http://openwetware.org/images/1/1c/TheBacterialPromoter.pdf
* http://en.wikipedia.org/wiki/Protecting_group#Carboxylic_acid_protecting_groups protecting groups
* http://en.wikipedia.org/wiki/Oligonucleotide_synthesis#Nucleoside_phosphoramidites
* http://en.wikibooks.org/wiki/Structural_Biochemistry/Nucleic_Acid/Nitrogenous_Bases/Purines

Latest revision as of 05:06, 23 August 2015

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