Oligonucleotides useful as adaptors in DNA cloning, adapted DNA molecules, and methods of preparing adaptors and adapted molecules

• US 4,321,365 A
• Filed: 10/19/1977
• Issued: 03/23/1982
• Est. Priority Date: 10/19/1977
• Status: Expired due to Term

• Current Assignee: National Research Council Canada
• Original Assignee: Research Corporation
• Inventors: Wu, Ray J., Narang, Saran A., Bahl, Chander P.
• Primary Examiner(s): Hazel, Blondel
• Application Number:
  US05/843,422
• Time in Patent Office: 1,616 Days
• Field of Search: 536/26-29, 435/317, 435/172
• US Class Current: 536/24.2
• CPC Class Codes: C07H 21/00 Compounds containing two or more mononucleotide units having separate phosphate or polyphosphate groups linked by saccharide radicals of nucleoside groups, e.g. nucleic acids
• CPC Class Codes: C12N 15/00 Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor mutants or genetically engineered microorganisms, per se C12N1/00, C12N5/00, C12N7/00; new plants per se A01H; plant reproduction by tissue culture techniques A01H4/00; new animals per se A01K67/00; use of medicinal preparations containing genetic material which is inserted into cells of the living body to treat genetic diseases, gene therapy A61K48/00

Abstract

Synthetic oligonucleotides have been designed and prepared which are useful in the molecular cloning of a variety of DNA molecules. By means of such oligonucleotides, genetic informational material, e.g., DNA, can be joined to a cloning vehicle and transferred into host cells by transformation. Additionally, a method for determining whether genetic informational material has been transferred into transformed host cells has been developed.

Claims

1. An adaptor molecule useful for attaching DNA containing genetic information to a cloning vehicle consisting of DNA which comprises a double-stranded oligodeoxyribonucleotide having at one end thereof a protruding nucleotide sequence which is the recognition site for a restriction endonuclease.

2. The adaptor molecule of claim 1 wherein the protruding nucleotide sequence is a part of the recognition site for EcoRI endonuclease.

3. The adaptor molecule of claim 1 wherein the protruding nucleotide sequence is a part of the recognition site for BamI endonuclease.

4. The adaptor molecule of claim 1 wherein the protruding nucleotide sequence is a part of the recognition site for HindIII endonuclease.

5. The adaptor molecule of claim 1 wherein the protruding nucleotide sequence is a part of the recognition site for PstI endonuclease.

6. The adaptor molecule of claim 1 wherein the protruding nucleotide sequence is a part of the recognition site for SalI endonuclease.

7. The adaptor molecule of claim 1 wherein the protruding nucleotide sequence is a part of the recognition site for HaeII endonuclease.

8. The adaptor molecule of claim 1 wherein the protruding nucleotide sequence is a part of the recognition site for XmaI endonuclease.

9. The adaptor molecule of claim 1 wherein the protruding nucleotide sequence is a part of the recognition site for BglII endonuclease.

14. An adapted DNA molecule which comprises double-stranded DNA containing genetic information to be attached to a cloning vehicle, a first adaptor molecule according to either claim 1 or claim 10 attached to one end of said DNA and a second adaptor molecule according to either claim 1 or claim 10 attached to the other end.

15. An adaptor molecule of claim 1 having the structure:

##STR9##

16. An adaptor molecule of claim 1 having the structure:

##STR10##

19. An adaptor molecule of claim 1 containing up to about sixteen deoxyribonucleotides.

10. An adaptor molecule useful for attaching DNA containing genetic information to a cloning vehicle consisting of DNA which comprises a double-stranded oligodeoxyribonucleotide having at opposite ends thereof first and second protruding nucleotide sequences which are recognition sites for restriction endonucleases.

11. The adaptor molecule of claim 10 wherein said first and second protruding nucleotide sequences are recognition sites for the same restriction endonuclease.

12. The adaptor molecule of claim 10 wherein said first and second protruding nucleotide sequences are recognition sites for different restriction endonucleases.

17. An adaptor molecule of claim 10 having the structure:

##STR11##

13. An adaptor molecule useful for attaching DNA containing genetic information to a cloning vehicle consisting of DNA which comprises a single-stranded oligodeoxyribonucleotide having nucleotide sequences which include the recognition sites for two different restriction endonucleases at opposite ends thereof.

18. An adaptor molecule of claim 13 having the structure:

##STR12##