# US 5,352,607 A · Molecular clone of a chitinase gene from vibrio parahemolyticus

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- Number: US 5,352,607 A
- Document ID: US5352607A
- Status: Granted
- Kind: A
- Application number: 07/876,894
- Filed: 1992-04-29
- Granted: 1994-10-04
- Earliest claimed: 1987-05-22
- Assignees: Louisiana State University and Agricultural College
- Inventors: Roger A. Laine; Chin-Yih Ou; Jesse M. Jaynes
- Web page: https://patentgrep.com/patent/US5352607A

## CPC

- G01N 33/56961: {Plant cells or fungi}
- C12N 9/2442: {Chitinase (3.2.1.14)}
- C12Y 302/01014: Chitinase (3.2.1.14)
- Y10S 435/909: Vibrio
- G01N: INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES (measuring or testing apparatus or processes other than immunoassay, involving enzymes or microorganisms C12M, C12Q)
- C12N: MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA (microbiological testing media C12Q1/00)
- C12Y: ENZYMES
- Y10S: TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS

## Abstract

A process for cloning the chitinase gene of Vibrio parahemolyticus is provided, comprising the steps of cleaving the Vibrio parahemolyticus DNA with Sau3A, PSTI or other restriction enzyme, mixing the cleaved DNA fragments in the presence of pUC18 and T4 ligase to produce a composite plasmid, and inserting the composite plasmid in a DH5a strain of E. Coli.

## Claims

### Claim 1

A composite DNA molecule comprising:
(a) a fragment of cleaved Vibrio parahemolyticus DNA from pC139 containing a chitinase gene which expresses a chitinase of approximately 95 Kd or a fragment of said gene which produces an active chitinase; and
(b) a vector ligated to said fragment wherein said vector is a plasmid or phage.

### Claim 2 (depends on 1)

A composite DNA molecule according to claim 1, wherein said fragment has restriction endonuclease digestion sites as shown in FIG. 1.

### Claim 3 (depends on 2)

A composite DNA molecule according to claim 2, wherein said Vibrio parahemolyticus DNA is cleaved with Sau3A.

### Claim 4 (depends on 1)

A composite DNA molecule according to claim 1 wherein said vector is cleaved with BamHI prior to ligating to said fragment.

### Claim 5 (depends on 4)

A composite DNA molecule according to claim 4 wherein said vector is pUC18.

### Claim 6

An E. coli microorganism carrying a recombinant plasmid pC139.

### Claim 7

A strain of E. coli identified as NRRLB-18200.

### Claim 8

A composite DNA molecule containing a Vibrio parahemolyticus chitinase gene comprising:
(a) a fragment of cleaved Vibrio parahemolyticus DNA from pC139 containing a chitinase gene which expresses a chitinase of approximately 95 Kd; and
(b) a vector ligated to said fragment at the BamH-I site, wherein said vector is a plasmid or phage.