Fingerprinting and molecular phylogeny of some heterocystous cyanobacteria using ۱۶S rRNA, ITS regions and highly iterated palindromes as molecular markers

This study aimed to investigate phylogenetic relationships of cyanobacteria based on the ۱۶S rRNA, ITS genes and palindromic sequences of HIP, ERIC, and STRR. The use of Internal Transcribed Spacer (ITS) region secondary structures has been proposed for phylogenetic reconstructions. Sampling was done from five Ghanats of shallow aqueducts located at Gonbad Kavous villages (Golestan province, NE of Iran), and purified in Z۸ culture medium. After DNA extraction, ۱۶S rRNA and ITS genes were amplified and sequenced. The phylogenetic tree was created using the Likelihood Maximum method and the appropriate model with the help of Iqtree online web server. The secondary structure of ITS was drawn in different parts of helix D۱-D۱′, D۲, D۳, tRNAIle, tRNAAla, BOX B, BOX A, and V۳ using Mfold program. Then, phylogenetic analysis of fingerprints was converted to binary information with the presence and absence of separate, and reproducible bands in each DNA fingerprint pattern generated by PCR profiles of HIP, ERIC and STRR, and binary information was used to construct a composite dendrograms. The results showed that, the studied strains belonged to four families viz. Aphanizomenonaceae, Nostocaceae, Hapalosiphonaceae, and Calotrichaceae of subsections of order Nostocales. The results of the dendrograms clusters drawn from the proliferation of palindrome sequences confirmed the clustering of phylogenetic trees. However, the results of the variable sections found in sections D۱-D۱′ and Box-B of the ITS gene revealed unique secondary structures that did not have a similar pattern to their close counterparts. The overall results showed that, the data obtained from genomic fingerprints, in silico and phylogenetic analysis are very useful for distinguishing closely related strains of cyanobacteria. 

Keywords:

Aphanizomenonaceae , Calotrichaceae , Hapalosiphonaceae , in silico analysis , Nostocaceae