An integrated linkage map of perlemoen (haliotis midae)

Hepple, Juli-ann (2010-12)

Thesis (MSc (Genetics))--University of Stellenbosch, 2010.

Includes bibliography.

Title page: Dept. of Genetics, Faculty of Science.

Thesis

ENGLISH ABSTRACT: Haliotis midae, or Perlemoen, is the only cultured species of abalone in South Africa and is under great international demand. This species is considered endangered, making sustainable farming practises and law enforcement against poaching essential for maintaining wild stocks. A limited amount of broodstock animals are provided to each farm from which thousands of offspring are grown and exported. The prevention of inbreeding and preservation of genetic diversity within farmed stocks is necessary for future sustainable farming and production of genetically stable offspring. Further research into the genetic dynamics of Perlemoen will provide the knowledge for advanced management programs for optimal farming practises and essentially sustainable production. This study focuses on genetic linkage map development with the intention of future identification of markers associated with genes of economic importance, such as growth rate. Identification of markers linked to genes responsible for such phenotypic traits will ultimately allow farming practises to select naturally genetically superior animals for breeding, thereby enhancing production. For the construction of a genetic linkage map of H. midae, microsatellite markers were developed using two strategies: FIASCO and screening of next generation sequence-bysynthesis contig data. The FIASCO-derived markers were characterised by genotype screening in 32 individuals from a full-sib family and analysed using Mendelian segregation expectations. The Illumina-derived markers were characterised by genotype screening in 32 individuals from wild populations and analysed against Hardy-Weinberg expectations. Forty four microsatellite-family combinations were obtained from FIASCO of which 28 provided informative genotype results (32% success). Twenty two markers were developed from sequence-by-synthesis screening. Fourteen provided reliable genotypes (37%) and six conformed to Hardy-Weinberg expectations. These markers were used, in addition to 156 previously developed markers, to develop sex-specific and sex-average linkage maps in two full-sib families consisting of approximately 100 offspring each. One hundred and six polymorphic loci were used for linkage analysis (LOD>3) in both families. The number of linkage groups obtained from sex-specific maps ranged from 13-16. The average genome length ranged from 500 cM to 800 cM with an average marker spacing of 10 cM. The sex-average linkage map provided 18 linkage groups with an average genome length calculation of 1800 cM and average marker spacing of approximately 13 cM. The linkage maps created in this study are preliminary but provide a stepping stone towards a high density map incorporating high throughput markers. This also provides a base for QTL mapping studies, in which phenotypic traits of interest can be identified and associated to specific locations in the H. midae genome for marker-assisted selection.

AFRIKAANSE OPSOMMING: Haliotis midae, ook bekend as Perlemoen, is in groot internasionale aanvraag en is ook die enigste klipkous spesie waarmee in Suid Afrika geboer word. Hierdie spesie word as bedreig beskou en daarom is volhoubare boerdery bedrywe en wetstoepassing teen stroping noodsaaklik om wilde populasies te beskerm. Elke perlemoenplaas word met ‘n beperkte aantal broeidiere verskaf, waarvan die nageslag dan gekweek en uitgevoer word. Voorkoming van inteling en handhawing van genetiese diversiteit binne gekweekte populasies is noodsaakllik vir toekomstige volhoubare kweking en produksie van ń geneties stabiele nageslag. Verdere ondersoeke na die genetiese dinamika van Perlemoen sal die nodige kennis verskaf om sodoende gevorderde bestuursprogramme te ontwikkel, wat tot optimale kweek praktyke en effektiewe volhoubare produksie sal lei. Hierdie studie fokus op die ontwikkeling van ‘n genetiese koppelingskaart met die voorneme om toekomstige merkers te identifiseer wat met gene van ekonomiese belang, soos byvoorbeeld groei tempo geassosieerd is. Identifisering van merkers wat vir sulke fenotipiese eienskappe verantwoordelik is sal sodoende toelaat dat boerdery praktyke kan selekteer vir diere vir verbeterde teling en produksie. Mikrosatelliet merkers is ontwikkel om die genetiese koppelingskaart saam te stel. Die volgende twee strategieë is benut: FIASCO en sifting van volgende generasie volgordebepaling-deur-sintese “contig” data. Die FIASCO-afgeleide merkers is gekarakteriseer deur genotipiese sifting in 32 individue van ‘n volsib familie en is deur Mendeliese segregasie verwagtinge ge-analiseer. Die Illumina-afgeleide merkers is gekarakteriseer deur genotipiese sifting in 32 individue van wilde populasies en is met Hardy-Weinberg ewewig ge-analiseer. Vier en veertig mikrosatelliet-familie kombinasies is deur FIASCO verky, waarvan 28 informatiewe genotipiese resultate gelewer het (32% sukses). Twee en twintig merkers is vanaf volgordebepaling-deur-sintese sifting ontwikkel. Veertien van hierdie merkers het betroubare genotipes (37%) verskaf en ses het aan Hardy-Weinberg verwagtinge voldoen. Hierbenewens is 156 voorheen ontwikkelde merkers gebruik om geslagspesifieke en geslagsgemiddelde koppelingskaarte in twee volsib families saam te stel. Hierdie volsib families het uit ń naslag van 100 elk bestaan. Een honderd en ses polimorfiese lokusse is vir koppelingsanalise gebruik, waar ‘n LOD waarde groter as drie statisties betekenisvol geag was. Die aantal koppelingsgroepe verkry van geslagspesifieke kaarte het tussen 13 en 16 gewissel. Die gemiddelde genoom lengte het van 500 cM tot 800 cM met ‘n gemiddelde merker spasiëring van 10 cM. Die geslagsgemiddelde koppelingskaart het 18 koppelingsgroepe gehad met ‘n gemiddelde genoom lengte berekening van 1800 cM en ‘n gemiddelde merker spasiëring van ongeveer 13 cM. Die koppelingskaarte wat in hierdie studie geskep is, is voorlopig en verskaf ‘n grondslag vir die ontwikkeling van ‘n hoër digtheidskaart, wat hoë deurset merkers inkorporeer. Dit verskaf ook ‘n basis vir kwantitatiewe kenmerk lokus karteringstudies. Hierdie karteringstudies kan fenotipiese eienskappe van belang identifiseer en assosieer met spesifieke posisies binne die H. midae genoom vir merker bemiddelde seleksie.

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