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. 2018 Mar 9;9:73. doi: 10.3389/fgene.2018.00073

Table 2.

Strategies used for optimization of next generation sequencing (NGS) microsatellite genotyping of Prochilodus costatus.

NGS Run Strategy# N° of primer pairs N° of DNA samples Cartridge¥ N° of filtered reads ∗∗N° of alleles
1 Multiplex 8, 4 and 2 5 1 M 13,691
Monoplex 8 829,631
2 Multiplex 8 192 15 M 608,390 30
3 Pseudo-multiplex 8 192 1 M 486,130 49
4 Pseudo-multiplex 8 96 1 M 646,223 48
5 Pseudo-multiplex 8 288 1 M 1,051,375 52
6 Pseudo-multiplex 8 384 1 M 991,337 53

Not applicate due to the low number of individuals. ∗∗Total number of alleles obtained for eight loci and variable numbers of individuals. #Reactions used for amplification. Multiplex, in which all primer pairs were used to amplify a single DNA sample in a single reaction followed by the incorporation of the index and NGS sequencing; Pseudo-multiplex, in which each primer pair was individually used to amplify a DNA target, followed by pooling of the amplicons of one individual (eight loci per individual) and a second PCR for index incorporation and MiSeq sequencing; Monoplex, in which DNA target amplification and index incorporation were performed individually for each DNA target and followed by MiSeq sequencing. ¥Type of cartridge used. 1 M is a MiSeq Reagent Nano Kit v2 (1 M reads) and 15 M is a MiSeq Reagent Kit v2 (15 M reads).