Nuclear Ribosomal RNA Gene (18S rDNA)
The aligned fragment of the COI gene was 503 bp long excluding the amplification primers, in total 253 individuals from fourteen stations were sequenced successfully(https://doi.org/10.1594/PANGAEA.909707 ). The alignment does not contain gaps, translation of sequences to amino acid sequences revealed no frameshifts mutations or stop codons. The analysis of the sequences identified 28 haplotypes, 70 sites were polymorphic and 56 parsimony informative. The haplotype network (Fig. 1) showed two sharply distinct groups separated by 50 mutational steps. Group A is distributed all along the Antarctic Peninsula and shows two dominant haplotypes, Group B is distributed in Weddell Sea, Potter Cove, Palmer Station, Paradise Bay, and Rothera Station, again with two common haplotypes that are mainly present in Palmer Station. In addition, there are 18 rare haplotypes, represented by one or two individuals from a single location.
The aligned sequences from the 18S fragment, containing the V4 ribosomal expansion segment, were 877 bp long. In total 312 individuals were sequenced, the alignment contained no gaps (https://doi.org/10.1594/PANGAEA.909707). We found 70 polymorphic sites, all parsimony informative, and after phase haplotype reconstruction, 10 haplotypes were recognized. From the 70 polymorphic informative sites, a single site at position 444 contained two variants that were congruent with the division among the mitochondrial groups A and B (Fig. 1). A single individual (collection code 291) showed both nucleotides (thymine and cytosine, respectively), this can be interpreted as 291 being heterozygous or hybrid (see discussion section 4.4).
The highest haplotype diversity for COI was in Potter Cove, but almost all the populations presented high values of diversity except for Livingston Island and the Scotia Sea. On the other hand, the highest haplotype diversity for 18S was in Burdwood Bank/MPA NamuncurĂ¡, but also Potter Cove, the Scotia Sea and, Shetland L45 and L46 presented high diversity values (Supplemental information, Table 1). Sequencing of COI was not possible for some individuals, several pairs of primers were tested (Bishop et al., 2013; Folmer, Hoeh, Black, & Vrijenhoek, 1994; Monniot, Dettai, Eleaume, Cruaud, & Ameziane, 2011) with no successful amplification, possible reasons for this are discussed later (section 4.1).