5% agarose gel electrophoresis The sulfotransferase cyrJ gene re

5% agarose gel electrophoresis. The sulfotransferase cyrJ gene required for tailoring reaction to complete the biosynthesis of the CYN was applied to assess the toxigenic potential of 24 water samples collected from Bytyńskie (BY) and Bnińskie (BN) lakes. The cyrJ gene was identified in 10 water samples from BY, and only two water samples collected at the beginning of the monitoring period in 2007 did not contain cyrJ gene (Table 2). However, in both samples, no CYN was found in the cells. In BN, the cyrJ gene was identified in all 12 water samples (Table 2). The presence of toxigenic cyanobacteria capable of producing cytotoxin throughout the season corresponded with the occurrence of CYN in 11

samples, with one exception, at the beginning of the monitoring period, that is, in the samples find more collected on 25 July 2007 (Table 2).

Summing up the cyrJ gene was detected in 22 of 24 investigated water samples. That observation indicated that the producers of CYN appear to be widespread in both lakes in the Western Poland (Table 2). Panobinostat manufacturer The PCR analysis of the water samples confirmed that cyrJ, which was originally recommended by Mihali et al. (2008) as a good candidate for determination of the toxin probe, can also be used for early detection of CYN-producing cyanobacteria in Polish lakes. In the study of Mihali et al. (2008), the screening of CYN-producing and nonproducing strains of C. raciborskii, Anabaena circinalis and Aph. ovalisporum revealed that the cyrJ sulfotransferase gene was present only in CYN-producing strains (Mihali et al., 2008). Mihali et al. (2008) emphasized that cyrJ gene is more specific than common cyanobacterial genes of NRPS (nonribosomal peptide synthetase) and PKS (polyketide synthase) and therefore can give fewer cross-reactions with other gene clusters. The results described, represent the first, to our best knowledge, genetic evidence for the occurrence of the CYN-producing cyanobacteria in Polish water bodies and the second, after German lakes, in the Central Europe. To identify

the source of cyrJ gene detected in our water samples, the PCR products from two samples from BY and two samples from BN, collected on 18 August 2006 and 30 August 2007, were subjected to cloning and sequencing. All the PCR products had the same nucleotide sequence. The blast homology search revealed that this sequence is in 99% similar Aprepitant to cyrJ gene of C. raciborskii and Aphanizomenon sp. However, all the sequenced samples carry the 6-nucleotide fragment, specific for cyrJ gene of Aphanizomenon sp., which is not present in relevant sequence in C. raciborskii genome (Fig. 1). Therefore, it may be concluded that all the PCR products were amplified based on cyrJ gene of Aphanizomenon sp. The activity of Aphanizomenon genus in the production of CYN was previously observed in the sample containing Aph. ovalisporum (pks/ps and cyrJ genes) or Anabaena bergii (pks/ps genes) obtained from Australian cultures (Schembri et al.

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