Fingerprinting and molecular phylogeny of some heterocystous cyanobacteria using 16S rRNA, ITS regions and highly iterated palindromes as molecular markers

Document Type : Research Paper

Authors

1 Assistant Prof., Department of Biotechnology, Science and Research Branch, Islamic Azad University, Tehran, Iran

2 Assistant Prof., Department of Genetics, Faculty of Advanced Science and Technology, Tehran Medical Sciences, Islamic Azad University, Tehran, Iran

10.22092/botany.2022.358594.1306

Abstract

This study aimed to investigate phylogenetic relationships of cyanobacteria based on the 16S 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 Z8 culture medium. After DNA extraction, 16S 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 D1-D1′, D2, D3, tRNAIle, tRNAAla, BOX B, BOX A, and V3 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 D1-D1′ 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


Article Title [فارسی]

انگشت‌نگاری و فیلوژنی مولکولی برخی از سیانوباکتری‌های هتروسیت‌دار با استفاده از مناطق 16S rRNA، ITS و پالیندروم‌های به ‌شدت تکراری به ‌عنوان مارکرهای مولکولی

Authors [فارسی]

  • بهاره نوروزی 1
  • حسین فهیمی 2
1 استادیار زیست‌شناسی مولکولی سیانوباکتری‌ها، گروه بیوتکنولوژی، دانشگاه آزاد اسلامی، واحد علوم و تحقیقات، تهران، ایران
2 استادیار گروه ژنتیک، دانشکده علوم و فناوری‌های نوین، دانشگاه علوم پزشکی آزاد اسلامی تهران، ایران
Abstract [فارسی]

هدف از این مطالعه، یافتن روابط فیلوژنتیک سیانوباکتری‌ها براساس ژن‌های 16S rRNA و ITS و توالی‌های پالیندرومی HIP، ERIC و STRR بوده است. به این منظور، نمونه‌برداری از پنج چشمه از قنات‌های کم‌عمق روستاهای گنبدکاووس (استان گلستان) انجام شد و در محیط کشت Z8 خالص‌سازی گردید. پس از استخراج DNA، ژن‌های 16S rRNA و ITS تکثیر و توالی‌یابی شدند. درخت فیلوژنتیک با استفاده از روش Likelihood Maximum و مدل مناسب به کمک وب سرور برخط Iqtree server ساخته شد. ساختار ثانویه ITS، در بخش‌های مختلف مارپیچ D1-D1′، D2، D3، tRNAIle، tRNAAla، BOX B، BOX A و V3 به کمک برنامه Mfold رسم شد. سپس، آنالیز فیلوژنتیک انگشت‌نگاری‌ها به کمک حضور و عدم حضور باندهای مجزا و قابل تکثیر در هر الگوی انگشت‌نگاری DNA تولید شده با پروفایل‌های PCR HIP، ERIC و STRR، به اطلاعات دوتایی تبدیل برای ساختن دندروگرام مرکب، استفاده شد. نتایج نشان داد که سویه‌های مورد بررسی متعلق به چهار تیره به اسامی Aphanizomenonaceae، Nostocaceae، Hapalosiphonaceae و Calotrichaceae از زیربخش راسته Nostocales بودند. نتایج کلاسترهای دندروگرام‌های رسم شده حاصل از تکثیر توالی‌های پالیندرومی تاییدکننده کلاستربندی‌های درخت‌های فیلوژنتیکی بود. به هر ‌حال، نتایج حاصل از بخش‌های متغیر در بخش‌های D1-D1′ و Box-B ژن ITS، ساختارهای ثانویه منحصر به ‌فردی یافت گردید که الگوی مشابهی با سویه‌های نزدیک به خود را نداشتند. نتایج کلی نشان داد که داده‌های حاصل از انگشت‌نگاری‌های ژنومیک، آنالیزهای درون‌رایانه‌ای و فیلوژنتیکی، برای تمایز سویه‌های نزدیک به هم سیانوباکتری‌ها بسیار مفید می‌باشند.
 

Keywords [فارسی]

  • آنالیزهای درون رایانه‌ای
  • Aphanizomenonaceae
  • Calotrichaceae
  • Hapalosiphonaceae
  •  Nostocaceae
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Volume 23, Issue 1 - Serial Number 63
September 2022
Pages 79-104
  • Receive Date: 06 May 2022
  • Revise Date: 17 July 2022
  • Accept Date: 19 July 2022
  • First Publish Date: 26 July 2022