Methods and Compositions for Production of Methane Gas
a technology of methane gas and compositions, applied in the field of methods and compositions for the production of methane gas, can solve the problems of explosive, sub>2 /sub>leaking through containers, and difficult handling
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example 1
Generation of Genetically Modified Cyanobacteria
The unicellular freshwater cyanobacterium Synechococcus elongates PCC7942 (previously known as Anacystis nidulans R2) and plasmid-containing derivatives were used in this study. Zymomonas Mobilis pyruvate decarboxylase gene (zmPDC) was cloned from plasmid pLOI295 (Ingram et al., Appl Environ Microbiol. 53(10): 2420-2425 (1987)). Pyruvate oxidase (POX) gene was cloned from the genomic DNA library of Lactobacillus plantarum (ATCC). The aldehyde dehydrogenase (aldDH) and acetate kinase (ackA) genes were cloned from Synechococcus elongates PCC 7942 genome directly.
Cells were grown in 200 BG-11 medium in 500 mL Erlenmeyer flask capped with BUGSTOPPER™ 10 (Whatman Inc., Clifton, N.J.). The cultures were agitated constantly at the speed of ˜100 rpm on a stirrer plate. The cultures were grown at either room temperature, or at 30° C. under white fluorescence light (1000-1200 lux). The growth was monitored by measuring the optical density at 730...
example 2
Production of Acetic Aced from Genetically Modified Cyanobacteria
To produce acetic acid, Synechococcus PCC7942 were first transformed with the pJX352 plasmid or the pJX373 plasmid and plated on BG11 / carbenicillin and incubated at room temperature under illumination for several days until single colonies developed. Single colonies were then picked using a sterile P200 pipette tip and inoculated 4 ml of liquid BG11 / carbenicillin growth medium in a glass tube. The culture was incubated with shaking at room temperate overnight under illumination. Prior to starting the large scale culture, the optical density at 730 nm of the 4 ml culture was measured when the culture reached a fairly green color. Once the optical density reached approximately 1.0 (stationary growth phase), 2 ml of the starter culture was transferred to a sterile 200 ml BG11 / carbinicillin liquid growth medium in a 500 ml flask. The 200 ml culture was grown at room temperate while shaking and under illumination. Periodica...
example 3
Growth of Mentanobacterium on Acetic Acid Containing Medium from Genetically Modified Cyanobacteria
Methanosarcina barkeri strain was obtained form the American Type Cell Culture (#29786). Methanosarcina medium was modified from ATCC media #1043 broth to include 0.0348% K2HPO4, 0.0227% KH2PO4, 0.05% NH4CL, 0.25% CaCl2, 0.00002% FeSO4.7H2O, 0.3% Trace Elements Solution SL-6, 1% Wolf's Vitamin Solution, 0.2% Yeast Extract, 0.2% Casitone, 0.225% NaCl, 0.0001% Resazurin, 2% NaHCO3 solution, and 2% Na2S / Cysteine reducing agent. All the ingredients were mixed except Wolf's Vitamin Solution and Na2S / Cysteine solution and autoclaved at 121° C. for 15 minutes. Wolf's Vitamin Solution and Na2S / Cysteine solution were subsequently added. The pH of the medium was adjusted to 6.8 before autoclave. All medium was prepared in an atmosphere of oxygen free 80% N2-20% CO2. All media were dispensed into Hungate tubes or flasks closed with butyl rubber stoppers and aluminum seals. The solution was then t...
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