Abstract
A study was conducted to construct a preliminary linkage map for molly, Poecilia and to test
association between markers and QTL traits. In I inkage map construction, three major steps were
involved. First is the development of mapping population. Background genetic information of the
parental species such as occurrence of triploidy and genetic variability were crucial before any
attempt to develop a mapping population. Triploid detection was carried out in this study by using
highly polymorphic microsatellite (SSRs) primer pairs. Ten SSRs primer pairs could clearly
differentiate among one, two and three alleles which could be used to infer different ploidy
levels. The presence of two alleles indicates a diploid at that particular locus and evidence of
three alleles could be inferred as triploid. If such association was direct, the occurrence of
triploidy in molly fish sampled in this study was extremely low and negligible. The genetic diversity
among the four species of live-bearer fishes namely Xiphophorus maculatus, Xiphophorus
helleri, Poecilia reticulata and Poecilia latipinna were revealed using 15 polymorphic SSRs primer
pairs. A total of 131 samples which comprised of these four species of live-bearers were collected
in the middle of October 2009 from Aquatic International, Subang Jaya, Malaysia. Results showed
that the clustering pattern of both Xiphophorus and Poecilia as revealed by these molecular markers seems to be in
accordance with their taxonomy. In P. latipinna, the occurrence of inbreeding or
outbreeding was absent as indicated by the level of heterozygosity. Majority of alleles were
homozygous within P. latipinna and this seems to resemble the genome characteristic of a
purebred fish. Purebred fish is an ideal parental material in any
mapping population.
A mapping population developed through backcross and testcross strategy was
successfully attempted. A parallel study on the mode of inheritance of background body colour had
also been initiated by evaluating body colour of progenies derived from
crosses between two different colour variants of molly, [P. latipinna (non-black) and P.
sphenops (black)]. Non-black body colour was found to be completely dominant over
black and was not sex-linked. Multiple genes interaction which acted nonadditively was also found
to be influencing this phenotypic trait. However, such interaction effect was
restricted in crosses generated between genetically related fish.
The second step in linkage map construction is identification of polymorphism using
molecular markers. A total of 142 SSRs primer pairs which includes those that were
developed for Xiphophorus maculatus (123 primer pairs) and Poecilia reticulata (19 primer pairs)
were screened and 95 primer pairs were able to be amplified on the DNA
of molly, Poecilia. However, only 29 SSRs primer pairs were polymorphic when genotyped on 77
progenies derived from two backcrosses (n = 56) and a testcross (n = 21). The fish samples used for
the development of mapping population were collected from Aquatic International Subang Jaya, Malaysia.
The final step is linkage analysis of polymorphic markers. Among the 29 polymorphic SSRs primer
pairs identified, only 18 were informative and could be used to construct the linkage map. The
constructed linkage map consists of four linkage groups covering a map size of 516.1 cM. One
potential QTL, dorsal fin length was successfully associated between marker regions ofMsd021, Msb069 and Msb068 on linkage group 2.
The major drawback found in this study was low percentage of polymorphic SSRs primer pairs. Such
condition not only greatly restricted the number of markers that could be positioned on the linkage
map but also minimizes the number of linkage group that could be generated. Map enrichment effort
using both amplified fragment length polymorphism (AFLP) markers and development of new SSRs primer
pairs for this species are strongly suggested.