Abstract: Fibrinogen is obtained from milk by contact with a cation exchange chromatography substrate under conditions where the fibrinogen binds to the substrate, followed by optional washing of the substrate. The bound fibrinogen is removed from the substrate by irrigation under increased ionic strength or increased pH conditions. The obtained fibrinogen may be naturally produced or transgenic.

Abstract: Peptides can be produced in and purified from the milk of transgenic animals. The peptides are made as fusion proteins with a suitable fusion partner such as &agr;-lactalbumin, which is a natural milk protein. The fusion partner protein acts to promote secretion of the peptides and, at least in the case of &agr;-lactalbumin, allows a single-step purification based on specific affinity. The peptide is released from the purified fusion protein by a simple cleavage step and purified away from the now liberated &agr;-lactalbumin by repeating the same affinity purification method. A particular advantage of producing peptides via this route, in addition to the obvious advantages of high yield and biocompatibility, is that specific post-translational modifications, such as carboxy terminal amidation, can be performed in the mammary gland.

Abstract: Peptides can be produced in and purified from the milk of transgenic animals. The peptides are made as fusion proteins with a suitable fusion partner such as &agr;-lactalbumin, which is a natural milk protein. The fusion partner protein acts to promote secretion of the peptides and, at least in the case of &agr;-lactalbumin, allows a single-step purification based on specific affinity. The peptide is released from the purified fusion protein by a simple cleavage step and purified away from the now liberated &agr;-lactalbumin by repeating the same affinity purification method. A particular advantage of producing peptides via this route, in addition to the obvious advantages of high yield and biocompatibility, is that specific post-translational modifications, such as carboxy terminal amidation, can be performed in the mammary gland.

Abstract: A method for purifying human or other alpha-1 proteinase inhibitor (&agr;1-PI) from a solution (which may be derived from the milk of a transgenic animal expressing the &agr;1-PI) which comprises contacting the solution with a cation exchange substrate under conditions sufficient to bind non-tg-&agr;1-PI contaminants to the substrate while not substantially binding tg &agr;1-PI to the substrate. Using the preferred embodiment, the purified tg &agr;1-PI contains as little as 40 pg non-&agr;1-PI-whey protein per mg total protein.

Abstract: Poor expression yields of recombinant human factor IX are attributable to aberrant splicing in heterologous expression systems such as transgenic hosts for example, transgenic nonhuman mammal The aberrant splicing sites have been identified as (a) a donor site including mRNA nucleotide 1085; and (b) an acceptor site including mRNA nucleotide 1547; adopting the mRNA nucleotide numbering of FIG. 2 of the drawings. Improved factor IX expression sequences have at least one of these sites engineered out, so as to prevent or reduce the effect of aberrant splicing and to increase yields. The improved DNA sequences may also be useful in gene therapy.

Abstract: Proteinaceous products can be produced by transgenic animals having genetic constructs integrated into their genome. The construct comprises a 5'-flanking sequence from a mammalian milk protein gene (such as beta-lactoglobulin) and DNA coding for a heterologous protein other than the milk protein (for example a serin protease such as alpha.sub.1 -antitrypsin or a blood factor such as Factor VIII or IX). The protein-coding DNA comprises at least one, but not all, of the introns naturally occurring in a gene coding for the heterologous protein. The 5'-flanking sequence is sufficient to drive expression of the heterologous protein.

Abstract: The present invention utilizes genetic engineering techniques to prepare non-human transgenic mammals that express human .alpha.-lactalbumin in their milk at a concentration of 2 mg/ml or greater. The invention also includes methods of preparing human .alpha.-lactalbumin in, for example, mice and cows. Also taught are methods for preparing human .alpha.-lactalbumin in which from one to four of its natural phenylalanine residues have been substituted by another amino acid.

Abstract: Proteinaceous products can be produced by transgenic animals having genetic constructs integrated into their genome. The construct comprises a 5'-flanking sequence from a mammalian milk protein gene (such as beta-lactoglobulin) and DNA coding for a heterologous protein other than the milk protein (for example a serin protease such as alpha.sub.1 -antitrypsin or a blood factor such as Factor VIII or IX). The protein-coding DNA comprises at least one, but not all, of the introns naturally occurring in a gene coding for the heterologous protein. The 5'-flanking sequence is sufficient to drive expression of the heterologous protein.