Alpha1-3 galactosyltransferase gene and promoter

• US 20030203427A1
• Filed: 04/19/2002
• Published: 10/30/2003
• Est. Priority Date: 10/22/1999
• Status: Abandoned Application

• Current Assignee: University of Pittsburgh of The Commonwealth System of Higher Education
• Original Assignee: University of Pittsburgh of The Commonwealth System of Higher Education
• Inventors: Koike, Chihiro
• Application Number:
  US10/125,994
• Publication Number:
  US 20030203427A1
• Time in Patent Office: Days
• US Class Current: 435/69.1
• CPC Class Codes: A01K 2217/05 Animals comprising random inserted nucleic acids (transgenic)
• CPC Class Codes: A01K 2217/075 inducing loss of function, i.e. knock out
• CPC Class Codes: C12N 9/1051 Hexosyltransferases (2.4.1)

Abstract

The present invention provides a recombinant expression cassette comprising an α1-3 galactosyltransferase promoter operably linked to a polynucleotide for expression. The invention also provides a recombinant mutating cassette comprising a region of homology to an α1-3 galactosyltransferase genomic sequence. The cassettes can be employed to express foreign genes or to disrupt the native α1-3 galactosyltransferase genomic sequence, particularly within an animal. Thus, the invention also provides transgenic animals and methods for their production and use.

Claims

1. A recombinant expression cassette comprising an α

1-3 galactosyltransferase promoter operably linked to a polynucleotide for expression, other than a polynucleotide encoding α

1-3 galactosyltransferase.

2. The recombinant expression cassette of claim 1, wherein said polynucleotide for expression encodes a protein.

3. The recombinant expression cassette of claim 2, wherein said protein is a fucosyltransferase, a galactosyltransferase, a β

-acetylgalactosaminyltransferase, an N-acetylglycosaminyltransferase, an N-acetylglucosaminyltransferase, a sialyltransferase, or a sulfotransferase.

4. The recombinant expression cassette of claim 2, wherein said protein is a Type I fucosyltransferase, a Type II fucosyltransferase, an α

2-3 sialyltransferase, or an α

2-6 sialyltransferase.

9. A vector comprising the recombinant cassette of claim 1.

10. A transgenic cell harboring the vector of claim 9.

13. A chromosome comprising the recombinant cassette of claim 1.

14. A transgenic cell harboring the chromosome of claim 13.

15. The transgenic cell of claim 14, wherein said α

1-3 galactosyltransferase promoter is native to said cell.

16. The transgenic cell of claim 14, wherein said polynucleotide for expression displaces a native polynucleotide encoding α

1-3 galactosyltransferase.

17. The transgenic cell of claim 14, which is an embryonic stem cell, an ovum, a primordial germ cell, a spermatozoon, or a zygote.

18. The transgenic cell of claim 14, which expresses said polynucleotide for expression.

19. The cell of claim 18, wherein said polynucleotide for expression encodes a Type I fucosyltransferase, a Type II fucosyltransferase, an α

2-3 sialyltransferase, or an α

2-6 sialyltransferase, and wherein said cell produces said protein.

20. The transgenic cell of claim 14, wherein said cell produces a heterogenic complement regulatory protein (CRP).

21. An embryo consisting essentially of transgenic cells according to claim 14

22. An organ consisting essentially of transgenic cells according to claim 14.

23. A transgenic animal consisting essentially of transgenic cells according to claim 14.

24. The transgenic animal of claim 23, which is a cattle, a mouse, a pig, a cat or a dog.

5. A recombinant mutating cassette comprising a first region of homology to an α

1-3 galactosyltransferase genomic sequence adjacent to either a second region of homology to said α

1-3 galactosyltransferase genomic sequence or a polynucleotide for insertion.

6. The recombinant mutating cassette of claim 5, comprising first and second regions of homology to an α

1-3 galactosyltransferase genomic sequence flanking a polynucleotide for insertion.

7. The recombinant mutating cassette of claim 5, wherein a region of homology is homologous to an exon, an intron, or a promoter of said α

1-3 galactosyltransferase genomic sequence.

8. The recombinant mutating cassette of claim 5, wherein said polynucleotide for insertion comprises an expression cassette.

11. A vector comprising the recombinant cassette of claim 5

12. A transgenic cell harboring the vector of claim 11.

25. A chromosome comprising the recombinant cassette of claim 5.

26. A transgenic cell harboring the chromosome of claim 25.

27. An embryo consisting essentially of transgenic cells according to claim 26

28. An organ consisting essentially of transgenic cells according to claim 26.

29. A transgenic animal consisting essentially of transgenic cells according to claim 26.

30. The transgenic animal of claim 29, which is a cattle, a mouse, a pig, a cat or a dog.

31. A transgenic knockout animal comprising a homozygous disruption in an endogenous α

1-3 galactosyltransferase gene, wherein said disruption prevents the expression of a functional α

1-3 galactosyltransferase protein.

32. The transgenic knockout animal of claim 31, wherein cells isolated from said knockout animal exhibit an increased time of survival in the presence of human serum relative to comparable cells isolated from an animal having a wild type α

1-3 galactosyltransferase gene.

33. The transgenic knockout animal of claim 31, wherein the insertion replaces DNA at the start of the coding region of said α

1-3 galactosyltransferase protein.

34. The transgenic knockout animal of claim 31, wherein the insertion replaces the promoter of said wild type α

1-3 galactosyltransferase gene.

35. The transgenic knockout animal of claim 31, which produces at least one human protein selected from the group of proteins consisting of α

1-3 galactosyltransferase, α

(1,2) fucosyltransferase, and complement regulatory proteins.

36. The transgenic knockout animal of claim 31, which is a pig.