CLONING AND SEQUENCE ANALYSIS OF THE EMBRYONIC ALPHA- AND BETA-TYPE GOAT GLOBIN GENES: EVOLUTIONARY ANALYSIS OF THE GOAT ALPHA AND BETA GLOBIN GENE CLUSTERS.

摘要

A major field of genetic research centers around experiments to help elucidate the mechanisms of gene expression and control. Towards that end, a variety of hemoglobin genes, both (alpha)- and (beta)-type, have been isolated, sequenced, and analyzed from the artiodactyl (goat). In this dissertation, I report the identification of an embryonic (alpha)-type globin gene, (zeta), and an embryonic (beta)-type globin gene, (epsilon)('IV), from the goat.; Clones containing two adult (alpha) globin genes of the goat have previously been identified and sequenced. Additional studies have; revealed the presence of an embryonic (alpha) globin gene termed (zeta). Sequence analysis of the gene shows that its coding sequence differs greatly from the adult (alpha). In particular at amino acid position 38, the goat (zeta) gene encodes an amino acid replacement that may confer a higher intrinsic O(,2) affinity to the (zeta) globin protein, insuring; a sufficient O(,2) supply for the developing goat embryo. The cloning and sequencing of this gene completes the (alpha) globin locus of the goat, composed of 5'-(zeta)-('I)(alpha)-('II)(alpha)-3'.; Evolutionary comparisons of the goat (alpha) locus to other amphibian, avian, and mammalian loci reveal several interesting features. The divergence rates of specific (alpha)-globin genes have been analyzed and found to form an essentially straight line, concurrent with the neutralist views of evolution.; In the goat (beta) globin locus I have identified an embryonic gene, named (epsilon)('IV). By sequence homology to the human (psi)h gene, it appears that the (epsilon)('IV) gene is also a (eta)-type gene. Sequence analysis has; revealed a splice junction donor sequence mutation 5'-GT(A/G)AG- 3' (--->) 5'-GCGAG-3'; a lariat acceptor site mutation, 5'-PyXPyTPuAPy- 3' (--->) 5'-CTCTGGC-3', as well as various heme and subunit contact; amino acid defects in the resulting protein coded for by the (epsilon)('IV) gene. Thus, (epsilon)('IV) is most probably a nonfunctional (eta) gene. Complete analysis; of the six (epsilon) genes from the triplicated goat (beta) globin cluster, 5'-(epsilon)('I)(epsilon)('II)(beta)('X)(beta)('C)--(epsilon)('III)(epsilon)('IV)(beta)('Z)(beta)('A)--(epsilon)('V) (epsilon)('VI)(beta)('Y)(beta)('F)-3', has revealed that (epsilon)('I), (epsilon)('III) and (epsilon)('V); are derived from the (epsilon) gene of the primordial locus; and (epsilon)('II), (epsilon)('IV), and (epsilon)('VI) are derived from the (eta) gene of the primordial locus. Evolutionary analysis of the cluster has revealed 5' end gene conversion of the (eta) gene by the (epsilon) gene in the primordial locus, and (epsilon)('II) conversion of; (epsilon)('IV) sequences post-duplication. Comparison of coding sequence supports a long history of functional evolution for the (epsilon)('V) and (epsilon)('VI) genes, post-duplication, prior to becoming pseudogenes.