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Genetically engineered biological indicator

A biological indicator, genetic engineering technology, applied in the field of genetic engineering biological indicators, can solve problems such as impracticality

Active Publication Date: 2009-09-16
AMERICAN STERILIZER CO
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, as noted above, this is impractical for many users of items that need to be sterilized

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0156] Figure 4 The plasmid described in was constructed by restriction digestion of the base plasmid and ligation of the xylR regulator fragment to the base plasmid. After confirming the correct ligation of the xylR regulator to the base fragment, the process was repeated for the bgaB gene fragment and terminators T1 and T2 for this gene. When the genetic elements are fully assembled and proper assembly and orientation are confirmed, the plasmid is inserted into the host organism and may be referred to as plasmid #1.

Embodiment 2

[0158] Plasmids can be internalized using transformation techniques in host organisms using several methods. These methods may include protoplasts, electroporation, ballistic methods, induction of competence, transduction or chemical methods (calcium chloride). For this example, competent E. coli were transformed by chemical transformation methods. E. coli lacking any restriction system was stored at -70°C. Thaw E. coli on ice and mix. E. coli was exposed to plasmid #1 for 30 minutes on ice. The resulting E. coli / plasmid mixture was heat-shocked at 42°C for 30 seconds and transferred to an ice bath for 2 minutes. Warm medium was then added to the E. coli / plasmid mixture and incubated at 37°C for 60 minutes. Place samples on agar plates containing antibiotics and X-gal. Clones appearing blue were successfully transformed.

Embodiment 3

[0160] Bacillus subtilis was grown overnight in LB broth containing 0.5M sorbitol, washed three times in 10 wt% glycerol with 0.5M sorbitol and 0.5M mannitol, and allowed to grow in the presence of plasmid #1 subjected to electroporation. Electroporation was performed via a range of voltages (1800-2500V). Resulting organisms were recovered in medium (LB broth medium with 0.5M sorbitol and 0.5M mannitol) and recovered for three hours at 37°C. After recovery, organisms were plated on LB agar plates containing antibiotics for selection of transformed organisms and incubated at 37°C. Clones exhibiting growth were plated on LB agar plates containing inducer (2 wt% xylose), selectable marker (chloramphenicol at 5 mg / mL), and X-gal (80 mg / mL). Colonies that appeared blue on these agar plates were successfully transformed.

[0161] The transformed Bacillus subtilis was sporulated using the method described below. Spread an aliquot of vegetative Bacillus subtilis on an agar plate. ...

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Abstract

The disclosed technology relates to a genetically engineered biological indicator, comprising: at least one test organism and at least one reporter gene suitable for producing an indicator enzyme, the reporter gene being taken up by the test organism; and at least one repressor gene that inhibits expression of the reporter gene until the reporter gene is exposed to at least one inducer. A process and an apparatus for using the biological indicator are disclosed.

Description

technical field [0001] The technology disclosed in the present invention relates to genetically engineered biological indicators. The biological indicators can be used to monitor the effectiveness of one or more sterilization processes. Background technique [0002] Primarily in the healthcare industry, but also in many other commercial and industrial applications, it is often necessary to monitor the effectiveness of processes used to sterilize equipment, such as medical and non-medical devices, instruments, and other items and materials. The inclusion of sterilization indicators in the batches of items to be sterilized is often standard practice in these sterilization processes. This allows a straightforward method for analyzing the lethality of the sterilization process. [0003] Typical methods of sterility assurance generally involve exposing a sterilization indicator containing one or more test organisms (test organisms) to the sterilization process and then detectin...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/04C12Q1/68C07H21/04
CPCC12N9/2471C12N9/2468C12N15/75C12Y302/01023Y02A50/30
Inventor 菲利普·P.·弗朗西斯克维赫威廉·A.·伊拉瓦特里克拉·A.·克雷格
Owner AMERICAN STERILIZER CO
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