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Biomedical subjects

A L Archibald

Publications and source records attributed to A L Archibald.

At least 55 records · Page 3Linked to original sources
PubMed1994

Characterization of microsatellites from flow-sorted porcine chromosome 13.

Porcine flow-sorted Chromosome (Chr) 13 was PCR amplified with primers based on porcine short interspersed element (SINE) sequences. The product was cloned, gridded in microtiter plates, and screened with a [GT]10 oligonucleotide which gave 45 positive clones. Sequencing of these clones showed that 36 were unique, and 26 [GT]n microsatellites were characterized. Six other simple repeat sequences, the majority of which were associated with the 3' end of the SINE sequence, were also detected. Twenty-one primers sets were selected, and 13 of these detected useful polymorphisms in the grandparents (n = 26) of the European porcine mapping collaboration (PiGMaP) reference families. These 13 markers were mapped in the "PiGMaP" reference families, and a two-point linkage analysis was performed. The Lod scores indicated that three of the markers were not linked and the remaining 11 formed two linkage groups of two and nine markers respectively. The larger linkage group was also linked to the transferrin locus, permitting assignment of nine markers to porcine Chr 13.

Animals↗
PubMed1994

Mapping of the pig genome.

The past year has seen major developments in both the genetic (linkage) and physical maps of the porcine genome. Landmark loci have been established on all pig chromosomes and outline genetic maps have been elaborated. The use of pigs as models of human genetic diseases is likely to expand through the use of transgenic pigs.

Animals↗
PubMed1994

RFLP and linkage analysis of the porcine casein loci--CASAS1, CASAS2, CASB and CASK.

Restriction fragment length polymorphisms (RFLPs) were revealed at the porcine casein loci with the following combinations of restriction endonucleases and porcine cDNA clones: alpha s1-casein (TaqI); alpha s2-casein (BamHI); and beta-casein (SacI). These RFLPs were shown to be under simple monogenic control by segregation analysis of two- and three-generation families. The CASAS1, CASAS2 and CASB casein loci were also shown to be linked with no recombinant haplotypes observed amongst 77 meioses in Large White and Meishan F1 and F2 crosses. No recombinants were observed in a further 106 meioses that were informative for linkage between CASAS1 and CASAS2.

Alleles↗
PubMed1993

Enhancing the efficiency of transgene expression.

Two strategies for enhancing gene expression in transgenic animals are described with particular reference to targeting expression to the mammary gland. Gene constructs in which the protein-encoding DNA sequences are contained within a genomic segment (comprising most or all of the natural introns of the corresponding gene) are shown to be expressed more efficiently than their intronless counterparts. Secondly, co-integrating an otherwise poorly expressed transgene in the vicinity of an actively expressed transgene can dramatically improve its efficiency of expression.

Animals↗
PubMed1993

5' and 3' SINE-PCR allows genotyping of pig families without cloning and sequencing steps.

A microsatellite-containing clone, isolated from a pig Chromosome (Chr) 1-specific library was characterized by sequencing and computer analysis. The (CA)17 microsatellite motif was located at the 3' end of a short interspersed element (SINE) sequence at the position normally occupied by the oligo (A) stretch. Further computer analysis indicated that 12% of published pig SINE sequences contain dinucleotide repeat motifs adjacent to their 3' ends. By performing PCR with a single SINE primer in combination with a panel of arbitrarily selected unique primers, we have demonstrated that, as in human, polymorphisms can be detected and typed in pig family DNAs. A large number of SINE primer x unique primer combinations have been screened for the ability to detect polymorphisms in pig reference family DNAs. This approach does not require prior sequence information other than that of the pig SINE. We have also found polymorphisms at the 5' ends of pig SINE sequences by similar methods, but with a primer facing out to the 5' end of the SINE.

Animals↗
PubMed1992

The role of the skeletal muscle ryanodine receptor gene in malignant hyperthermia.

Malignant hyperthermia (MH) is an inherited, potentially lethal condition in which sustained muscle contracture with attendant hypermetabolism and hyperthermia is triggered in humans, heterozygous for the gene defect, by inhalational anaesthetics and skeletal muscle relaxants, and in pigs, homozygous for the defect, by stress. Because muscle contracture could result from a defective Ca2+ release channel, we have focussed our attention on the linkage of MH to defects in the gene (RYR1) encoding the skeletal muscle Ca2+ release channel. We have cloned and sequenced human RYR1 cDNA and found restriction fragment length polymorphisms (RFLPs) in the human gene. We also localized RYR1 to human chromosome 19q13.1. Studies of the cosegregation of MH with these RFLPs established RYR1/MH linkage on human chromosome 19q13.1 (lod score of 4.2; recombinant fraction 0.0). We then sequenced MH and normal porcine RYR1 cDNAs. Mutation of C1843 to T, leading to substitution of Cys for Arg615, was the sole amino acid change noted between MH and normal animals. Linkage of this mutation to MH was established in a study of 338 informative meioses (lod score of 102; recombinant fraction 0.0). We identified the corresponding mutation in 1 of 35 human MH families studied and found cosegregation of the mutation and MH. The combination of a high lod score with crossing of a species barrier supports the causal nature of this mutation. Future studies are aimed at finding the major human MH mutations and establishing assays for their accurate diagnosis.

Amino Acid Sequence↗
PubMed1991

Production of pharmaceutical proteins in milk.

There is every reason to expect that it will be possible within the next few years to begin to use farm animals to produce large quantities of some of the human proteins that are needed for the treatment of disease. Revolutionary new opportunities for the production of novel proteins in milk have been created by the development of methods for gene transfer. Exploitation of these opportunities depends upon selection and cloning of milk protein genes and identification of the sequences that govern tissue specific hormonally induced expression in the mammary gland. Studies with three genes, ovine beta-lactoglobulin, rat beta-casein and whey acidic protein of rat and mouse, suggest that they may all meet this requirement. Fragments of the ovine beta-lactoglobulin, murine whey acidic protein and rabbit beta-casein genes have directed production of novel proteins in the milk of transgenic mice, sheep, rabbits and pigs. The proteins were biologically active and usually co-migrated with authentic proteins. In early experiments, protein concentration was low, but our recent observations suggest that fusion genes containing genomic clones direct production of concentrations of protein that are suitable for commercial exploitation. In the longer term, two approaches may offer the potential of more reliable expression. Control elements capable of directing expression that is independent of site of insertion of the gene, but dependent on the number of copies of the gene, have been identified for a small number of genes. The availability of such elements for the milk protein genes would increase the reliability of gene expression considerably. Alternatively, targeted mutation of genes may allow the insertion of coding sequences within an existing gene so avoiding position effects.

Animals↗