Inoculating preheated trays containing nutrient agar with photobacteria provides a means for mass culture of aerobic microorganisms in order to obtain large quantities of luciferase. To determine optimum harvest time, growth can be monitored by automated light-detection instrumentation

Chappelle, E. W.

Picciolo, G. L.

Rich, E., Jr.

English

NASA Technical Reports Server

.August 1969
	 Brief 69-10294 
NASA TECH BRIEF 
NASA Tech Briefs are issued to summarize specific innovations derived from the U.S. space program, 
to encourage their commercial application. Copies are available to the public at 15 cents each 
from the Clearinghouse for Federal Scientific and Technical Information, Springfield, Virginia 22151. 
Mass Culture of Photobacteria to Obtain Luciferase 
The problem: 
Devise a laboratory method for mass culture of aero-
bic microorganisms, particularly photobacteria, to ob-
tain large quantities of luciferase. 
The solution: 
Inoculate large preheated trays filled with nutrient 
agar with photobacteria and grow these microorga-
nisms in a controlled environment. Monitor growth by 
automated light-detection instrumentation to deter-
mine optimum harvest time. Selectively scrape the 
luminescent organisms from the agar surface with a 
rubber spatula in the dark and process the cell paste 
obtained to prepare luciferase. 
How it's done: 
Five strains of photobacteria are used, namely, 
Photobacterium fischeri. P. harveyi, P. pieranlonhi, 
P. phosphoreum, and Vibrio albensis. Stocks are rou-
tineIN cultured on a minimal medium.
(Ofltiflued Overleaf) 
fhis document was prepared under the sponsorship of the National
	 Government assumes any liability resulting from the use of the 
Aeronautics and Space Administration. Neither the United States	 information contained in this document, or warrants that such use 
Government nor any person acting on behalf of the United States
	 will be free from privately owned rights.
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Warm sterilized fiberglass trays, 50 x 75 x 4.5 cm, 
are filled to a depth of one centimeter with three 
liters of hot sterile agar medium, covered loosely with 
aluminum foil, and allowed to cool on a level surface. 
One hundred milliliters of a dense, luminescing culture 
are poured on the cooled agar tray and spread thinly 
and evenly with a bent glass rod or cotton swab. The 
tray is placed into the facilities used for temperature 
and humidity control. 
A polymethyl methacrylate plate bridges the agar 
surface to support a light sensor. Covering the entire 
tray with heavy-duty aluminum foil keeps the surface 
dark and minimizes evaporation and contamination 
during growth. A commercially available photomulti-
plier in a photometer monitors changes in lumines-
cence. The photomultiplier output feeds into a log-
arithmic amplifier to record the wide range of light 
produced over maximum growth. This amplifier com-
presses seven decades of response into a 0 to 10-volt 
output signal recorded on a strip-chart recorder with a 
mechanical timer set for recording 5 seconds of each 
IS-minute period. This allows continuous monitoring 
over any desired time period. The photograph shows 
the entire apparatus.
Trays are harvested at maximum luminescence in 
the dark to facilitate selection of the most luminous 
areas for scraping with a rubber spatula. Three liters 
of medium can yield as much as 100 grams of Photo-
bacterium-cell paste, which can be processed to pre-
pare enough luciferase for a large number of experi-
ments. 
Notes: 
I. Laboratories and small-scale pilot plants with lim-
ited facilities will find this method for mass culture 
of aerobic microorganisms very useful. Numerous 
biological components may be extracted from the 
microorganisms. 
2. Additional information may be found in a paper. 
"A Simple Method for the Mass Culture of Micro-
organisms with Reference to Photobacteria", by 
G.L. Picciolo, E.W. Chapelle, and E. Rich, Jr., 
Applied Microbiology 16, 954-955, June 1968. 
3. No further documentation is available. 
Patent status: 
No patent action is contemplated by NASA. 
Source: G. L. Picciolo, E. W. Chappelle 
and E. Rich, Jr. 
Goddard Space Flight Center 
-	 (GSC-10563) 
Brief 69-10294	 Category 04


Mass culture of photobacteria to obtain luciferase

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