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Based on these criteria, eight SNP-STR markers were finally selected and amplification refractory mutation system (ARMS-PCR) method was used to achieve allele-specific amplification.
Thus, 1,131 SNP markers were finally positioned on the F2 map.
The staining results of these 12 markers were finally evaluated based on the mean proportion of two EHCC cores in each case.
In addition, 36 markers with more than 12 missing data points had to be removed and hence 403 markers were finally used for map construction.
However, 446 of these markers were finally integrated into the map so that the loss of genetic information related to these markers was minimized.
A total of 38 markers were finally selected by combining quality score, power of discrimination (PD), and whole genome distribution (1 3 markers/chromosome) (Table 1; Figure 1).
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The expression marker was finally detected using FACSCalibur flow cytometer (Becton Dickson, San Jose, California).
A COS marker was finally defined as a sequence presenting one single unigene from Pinus and Picea within the same family.
Linkage group–specific datasets with fewer than 8000 markers each were finally loaded into JoinMap to identify locus order.
For each experiment cells (PBMCs, monocyte derived macrophages, murine splenocytes or cell lines MCF7, A431 and HEK293) were stained with respective markers and were finally suspended in 50 µl of buffer (cold PBS with 1% FCS and 0.05% sodium azide) in 0.6 ml microcentrifuge tubes.
The marker sequence were finally assigned to certain ancestral blocks according to the best match length, E value, good alignment consistency with adjacent loci, good alignment consistency among bin markers, good alignment consistency with other published genetic map of B. juncea.
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