240 PATENT ABSTRACTS

4508826 4 5 0 8 8 2 7

B A C T E R I O P H A G E D N A C L O N I N G V E C T O R T G I A N D

M I C R O O R G A N I S M S C O N T A I N I N G T G I

Forrest Foor, Gary Roberts assigned to Merck & Co tnc

M O L E C U L A R C L O N I N G V E C T O R S F O R U S E I N G R A M -

N E G A T I V E B A C T E R I A

Ronald Olscn assigned to Microlife Technics lnc

- Restriction Map of Phage TG1

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~ ' ~ m ~ M ~ ~ ~ I t ~ N ~ ~1 N~I~,.~ M

I!~I/17~,/I IW7 lff,~ / III!Y If II I Ill I II II Ill I IIII I I I II I I I

Disclosed is the novel bacteriophage TGI , TG1 derivatives, and the corresponding genome or nucleic acid components of such bacteriophages and derivatives of such genome, which are useful as DNA cloning vectors into organisms, such as bacteria, for example, Streptomyces cattleya N R R L 8057; portions of such phage genome are additionally useful as adjuncts in recombinant DNA cloning procedures, for example: (1) to permit the maintenance of cloned DNA in the host, either in an integrated state or as an auto- nomous element; (2) to serve as promoters for in- creasing expression of endogenous or foreign genes wherein said promoters are ligated to such genes or otherwise serve as promoters; and (3) to serve as regulatory elements for achieving con- trol over endogenous and foreign gene expres- sion; as cloning vectors, TGI , its deletion mutants, and other derivatives serve for the am- plification and transfer of DNA sequences (genes) coding for useful functions, for example, genes necessary for the production of the anti- biotic thienamycin, or genes necessary for the production of hepatitis B antigen, and of DNA sequences which are useful per se, for example, distinct plasmid vectors which are inherently useful; such modified cloning vectors (hybrid DNA molecules comprising the TG I genome or portions thereof and foreign DNA sequence)are introduced into the recipient organism by infec- tion, transfection or transformation; wherein the hybrid DNA functions in an integrated mode, in a lyric (vegetative phase) mode and/or in a plas- mid mode. Also disclosed are microorganisms comprising TG1 prophage and deletion and hybride (chimeric) derivatives thereof; and microorganisms comprising hybrid (chimeric) phage-plasmids and derivatives thereof.

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Improved cloning vectors derived from pROI614 are described. One of these vectors. pRO1727, is suitable for cloning using DNA cleaved with the restriction endonuclease, Pstl, and allows selection for the recovery of recom- binant plasmids using tetracycline resistance. The cloning efficiency observed for pRO1727 is higher than described previously for pRO1614 and the host range of this vector is now restricted to Pseudomonas bacteria. Another vector, designated pRO1729, is described and developed from pROI727 by deletion of a portion of its DNA and incorporation of a segment of DNA which encodes for resistance to the antibiotic, chloramphenicol. The chloramphenicol resistance determinant has a cleavage site for restriction endonuclease EcoRl within its chloramphenicol resistance determinant. Thus, DNA cloned into this site results in the loss of chloramphenicol resistance which can be detec- ted subsequent to a cloning experiment. Both pROI727 and pROI729 are more useful in Pseudomonas for cloning than pRO1614.

4510245

A D E N O V I R U S P R O M O T E R S Y S T E M

Lawrence Cousens, Graeme 1 Bell. Pablo D T Vatenzuela assigned to Chiron Corporation