Réaction #470399

ord-7983ec4dee9d4fc5808a36db902eeb1a

Équation de réaction

O=C[C@H](O)[C@@H](O)[C@@H](O)CO
L-arabinose
CC(C)S[C@@H]1O[C@H](CO)[C@H](O)[C@H](O)[C@H]1O
isopropyl-β-D-thiogalactopyranoside
CC1(C)S[C@@H]2[C@H](NC(=O)[C@H](N)c3ccccc3)C(=O)N2[C@H]1C(=O)O
ampicillin
CC1(C)S[C@@H]2[C@H](NC(=O)[C@H](N)c3ccccc3)C(=O)N2[C@H]1C(=O)O
ampicillin
→
OC[C@@H](O)[C@H](O)[C@@H](O)CO
xylitol
O=C[C@H](O)[C@@H](O)[C@@H](O)CO
L-arabinose

Réactifs

O=C[C@H](O)[C@@H](O)[C@@H](O)CO
L-arabinose
CC(C)S[C@@H]1O[C@H](CO)[C@H](O)[C@H](O)[C@H]1O
isopropyl-β-D-thiogalactopyranoside
—
( ZUC99/pATX112 )
CC1(C)S[C@@H]2[C@H](NC(=O)[C@H](N)c3ccccc3)C(=O)N2[C@H]1C(=O)O
ampicillin
CC1(C)S[C@@H]2[C@H](NC(=O)[C@H](N)c3ccccc3)C(=O)N2[C@H]1C(=O)O
ampicillin

Réactifs

Aucun

Conditions de réaction

Conditions détaillées
See reaction.notes.procedure_details.

Traitement

  1. 1
    workup.ADDITIONAfter incubation, an aliquot of the culture was diluted 100-fold into 20 mL of fresh LB media
  2. 2
    Autre(30° C., 250 RPM)
  3. 3
    Autrefor 2 hrs
  4. 4
    Autrewas incubated at 30° C.
  5. 5
    workup.STIRRINGwith shaking (250 RPM) and aliquots
  6. 6
    Autrewere removed at various time points after the IPTG induction

Mode opératoire

To test the activity of the XR/Lad1/Alx1 operon, plasmid pATX112 was inserted into strain ZUC99 by electroporation (ZUC99/pATX112). The strain was inoculated from a single colony into 3 mL LB media supplemented with ampicillin (200 mg/L) and incubated overnight at 30° C. with shaking (250 RPM). After incubation, an aliquot of the culture was diluted 100-fold into 20 mL of fresh LB media contain L-arabinose (1% w/v) and ampicillin (200 mg/L) and incubated (30° C., 250 RPM) for 2 hrs. The culture was induced with isopropyl-β-D-thiogalactopyranoside (IPTG, 1 mM). The culture was incubated at 30° C. with shaking (250 RPM) and aliquots were removed at various time points after the IPTG induction. The samples were monitored for xylitol formation from L-arabinose by HPLC analysis using an Aminex HPX-87P column (BioRad, USA). After 24 hrs post induction, the XR/Lad1/Alx1 operon strain (ZUC99/pATX112) displayed a 2% conversion of L-arabinose to xylitol, showing that the recombinant bacterium acquires the ability to produce xylitol from L-arabinose (Table 6).

Source

DOI: 10.6084/m9.figshare.5104873.v1Brevet: US08367346B2uspto-grants-2013_02