| Amplified Fragment Length Polymorphism (AFLP) markers were used to construct the linkage map of Fenneropenaeus chinensis. We sampled maternal individual from the fourth successively selected generation of disease resistance population of F. chinensis and paternal individual from wild population, respectively. Artificial insemination was used to generate F1 family; F2 families totally 42 bodies came from sisterhood intercross between F1 families. The F2 progenies were firstly subjected to the WSSV challenge, the survival time were recorded as the index of disease resistance traits of F. chinensis, the data of its total length, body length and weight were recorded as well when the individual was found death. All the data recorded would be involved in the QTLs mapping. Sixty-two primer combinations produced 529 segregating bands; of which 253 segregate in a 1:1 model and 276 in a 3:1 model. The female and male 1:1 ratio makers were used to construct the respective linkage based on the pseudo-testcross strategy. The 3:1 ratio segregant markers were used to construct the common linkage map of F. chinensis using F2 intercross model. The female linkage map contains 94 segregating markers, which were linked in 31 linkage groups (including linkage couples), covering 51.39% of F. chinensis genome with the interval of 12.20cM. The length of maximum linkage group was 82.9 cM, the maximum number of markers in a linkage group was 6. The male linkage map contains 81 segregating markers, which were linked in 25 linkage groups, covering 50.21% of F. chinensis with the interval of 11.45cM. The length of maximum linkage group was 103.8 cM, the maximum number of markers in a linkage group was 8. The 276 markers in a 3:1 model were used to construct the common F. chinensis linkage map based on F2 intercross model strategy. The common linkage map consists of 44 linkage maps with 129 markers, covering 48.08% of F. chinensis genome with the interval of 11.12cM. The length of maximum linkage group was 80.6 cM, the maximum number of markers in a linkage group was 8. All the linked markers distributed evenly in linage groups, no cluster marker was found in any linkage group. The density and coverage of these three linkage groups showed that they matched the medium density linkage map standard. The linkage mapping strategies we applied in F. chinensis and mapping families were discussed. Based on the three linkage maps, QTLs (Quantitative Trait Loci) associated with body length, total length, weight and disease resistance were analyzed by MAPMAKER/QTL1.1 software using interval mapping (IM). There were totally 11 loci were detected on 6 linkage groups. Four of them were associated with body length, two with total length, two with weight and three with disease resistance. The four QTLs loci associated with body length distributed on four different linkage groups, two of them linked with QTLs of total length, one of them linked with QTLs of weight, and the last one distributed in a single linkage group, which suggested that the trait of total length was polygenic controlled. The three QTLs with disease resistance distributed on three different linkage groups, two of them linked with two QTLs of weight, which suggested that the individual's disease resistance was related with the body's weight and multi-genes controlled the disease resistance as well. Except four QTLs additive value were negative, the other seven were positive. The dominance value only owned by the QTLs that based on the common linkage groups of F. chinensis and they were all positive. The variance explained ratio of 11 QTLs was from 23.4% to 66.9%. Following with the QTLs additive and dominant values, the individual's trait values of different genotypes were analyzed. The reasons that resulted in some very high variance explained were discussed based on the QTL mapping method, data recording and the size of mapping family. AFLP technique has been used in this study for detection the sex-specific markers of bluegill sunfish based on sex-type DNA pool strategy. The phenotypically female and male individuals that composed of sex-type genetic DNA pool were subjected to gonad anatomy to confirm their sexualities. Seven sex-specific markers were identified from 64 different primer combinations amplification across the female and male genomic DNA pools that composed of 24 female and male individuals, respectively. Six were female and one was male related marker. The sex-specific markers were subsequently analyzed individually: there were 16.67%-41.67% of 24 female or male individuals were detected sharing the sex-specific markers. In the linkage analysis, paternal and maternal bodies that selected from two different bluegill population that kept in South centers for several generations consisted in a single-pair parent. The single-pair parent and their 90 progenies were analyzed using as mapping family based on double-pseudo testcross strategy, sixty-four primer combinations that as same as in the sex-specific markers analysis produced totally 438 segregant loci, including 1 sex-specific and 9 co-dominant markers. In maternal linkage map, 192 markers distributed on 39 linkage groups (LOD≥4.0); There were 21 linkage groups that owned at least 4 markers, the maximum length and markers number of linkage groups were 122.9 cM and 14, respectively; the average markers number in each linkage group were 4.92, the average markers space was 11.29 cM, the genome coverage of maternal linkage map was 64.04%. In maternal linkage map, we have acquired a sex-specific linkage group 8. The cluster markers on linkage groups 5, 6, 8, 10 and 11 were tended to distribute by the end of linkage groups. For the paternal linkage map, 191 linked markers, including 4 co-dominant, distributed on 40 linkage groups (LOD≥4.0). The linkage groups that owned at least 4 markers number was 21, the average markers number of each linkage group was 4.77 in paternal linkage map, and average markers space is 10.58cM. The maximum length and markers number of linkage groups in paternal linkage map were 345.3cM and 19, respectively. The genome coverage of paternal linkage map was 66.26%. The linkage groups 11, 31 and 32 were found had cluster markers at the end of linkage groups. Bluegill sunfish karyotype showed that this kind of fish had diploid numbers of 48, consisting of telocentrics and acrocentrics with very short arms, in the linkage map of bluegill sunfish, many of the cluster AFLP markers distributed by the end of linkage groups, which were consistent with the chromosomes structure, but only the chromosomes become defined in bluegill sunfish, the locations of the cluster markers could be better defined. The maternal and paternal linkage group numbers were more than the haploid chromosomes number (2n=48) of bluegill, which suggested some linkage groups shared the same one chromosome, with supplement more AFLP markers and some other types marker, such as SSR, the number of linkage groups should match haploid chromosome number. The length of maternal linkage map was 129cM more than the paternal map, which was similar as in the other vertebrate organisms and other fishes, such as in rainbow trout and zebrafish. The maximum length of linkage groups in paternal map was 345.3 cM, we could not find any other matchable groups in maternal linkage map, which suggested that the difference between male and female chromosome of bluegill sunfish was obviously. Of all the 7 sex-specific markers, 6 were female specific and one was male, the chance that female owned heteromorphic sex chromosome or the female had one more chromosome than the male were good, namely the sex chromosome system of bluegill sunfish were ZW/WW or XX/XO type. The evidence that the paternal owned a much bigger linkage groups than the maternal, combined with the evidence that the green sunfish, its sex chromosome system was XX/XO, could hybridize with bluegill sunfish very easily, more suggested that the bluegill sunfish had XX/XO sex chromosome system. This work was the first study of linkage mapping of bluegill sunfish using AFLP markers, and it was the first time we tried to clear the sex determination mechanism from DNA markers and linkage mapping.
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