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18S rRNA and Its Use in Fungal Diversity Analysis

18S ribosomal RNA (18S rRNA) is a component of the small eukaryotic ribosomal subunit (40S), and 40S and 60S constitute eukaryotic ribosomes. As a structural RNA for<br>eukaryotic ribosomes, 18S rRNA, the homologue of 16S rRNA in prokaryotes and mitochondria, is thus one of the essential components of all eukaryotic cells. 18S rRNA genes that code for 18S rRNA are widely used in phylogenetic analysis and<br>environmental biodiversity screening.

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18S rRNA and Its Use in Fungal Diversity Analysis

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  1. 18S rRNA and Its Use in Fungal Diversity Analysis What is 18S rRNA? 18S ribosomal RNA (18S rRNA) is a component of the small eukaryotic ribosomal subunit (40S), and 40S and 60S constitute eukaryotic ribosomes. As a structural RNA for eukaryotic ribosomes, 18S rRNA, the homologue of 16S rRNA in prokaryotes and mitochondria, is thus one of the essential components of all eukaryotic cells. 18S rRNA genes that code for 18S rRNA are widely used in phylogenetic analysis and environmental biodiversity screening. Figure 1. Prokaryotic 70S ribosome and eukaryotic 80S ribosome. 18S rRNA as a marker for biodiversity studies 18S rRNA gene is a common molecular marker for biodiversity studies since it is highly conserved intra-species (similarities close to 100%) and assist in species-level analyses. Similar to 16S rRNA, 18S rRNA gene has nine variable regions (V1-V9). Previous studies tested the taxonomic resolutions of 18S rRNA gene at different taxonomic levels (Wu et al. 2015) come to the following conclusions: i. full-length 18S rRNA sequences or partial regions (around V2, V4, and V9) of the 18S rRNA gene are useful for the discrimination

  2. of samples at both the family and order levels; ii. V9 has a higher resolution at the genus level; iii. V4 is the most divergent region in length, which would be a marker candidate for the phylogenetic study of Acartia speicies. Once we obtained 18S rRNA sequences, they could be used for taxonomic resolutions and diversity analysis in eukaryotic communities. While 16S rRNA gene sequencing provides insight into bacterial diversity, 18S rRNA gene sequencing can offer insight into fungal diversity. Taxonomic structure of prokaryotic and eukaryotic microbial communities can be determined via 16S and 18S rRNA gene sequencing. It is crucial to understand the ecological niches that contribute to environmental pathogen development. What is the difference between 18S rRNA and ITS in metagenomic analysis? ITS (internal transcribed spacer region) is located between the 18S and 5.8S rRNA genes and has a high degree of sequence variation. Similar to 18S rRNA, ITS is often used in metagenomic analysis. However, 18S rRNA is mainly used for high resolution taxonomic studies of fungi, while the ITS region is mainly used for fungal diversity studies as a fungal barcode marker. Compared to 18S, ITS is more variable and thereby, more suitable as the genetic marker to measure intraspecific genetic diversity. Figure 2. Schematic diagram of the eukaryotic rRNA genes. Primers for 18S rRNA There are many available primers for 18S rRNA as shown in Table 1. With primers NS1 and NS8, we can obtain a greater length than 1,600 bp (the full-length of 18S rRNA is approximately 1800 bp). Alternatively, 18S rRNA sequences available in databases such as Silva (https://www.arb-silva.de/) and EukRef (http://eukref.org/) can be used for primer design. Table 1. Primers for 18S rRNA (from Berkeley university of California). Name NS1 CNS1 Primer Sequence GTAGTCATATGCTTGTCTC GAGACAAGCATATGACTACTG Tm 49 55

  3. NS2 NS3 NS4 NS5 NS6 NS7 NS8 TW9 TW10 TW11 TW12 CTW12 NS17 NS18 NS19 NS20 NS21-ag NS21-ls NS22 NS23 NS24 NS25 CNS25 NS26 CNS26 VANS1 MB1 CMB1 MB2 Basid 1 Basid 2 Basid 3 Basid 4 NS1.5R NS2.8R CNS2.8R AATTTGCGCGCCTGCTGCAA NS3.2R CGTATATTAAAATTGTTGAC CNS3.3R GACTACGAGCTTTTTAACGT CNS3.5R TTTCGCAGTAGTTTGTCTTA NS3.6R CAAACTACTGCGAAAGCATC CNS3.6R AATGAAGTCATCCTTGGCAG GGCTGCTGGCACCAGACTTGC GCAAGTCTGGTGCCAGCAGCC CTTCCGTCAATTCCTTTAAG AACTTAAAGGAATTGACGGAAG GCATCACAGACCTGTTATTGCCTC GAGGCAATAACAGGTCTGTGATGC TCCGCAGGTTCACCTACGGA TAAGCCATGCATGTCT GCGGTAATTCCAGCTCC GGAGTGGAGCCTGCGGCT AAGTCGTAACAAGGTTT AAACCTTGTTACGACTT CATGTCTAAGTTTAAGCAA CTCATTCCAATTACAAGACC CCGGAGAAGGAGCCTGAGAAAC CGTCCCTATTAATCATTACG GAATAATAGAATAGGACG AATATACGCTATTGGAGCTGG AATTAAGCAGACAAATCACT GACTCAACACGGGAAACTC AAACCTTGTTACGACTTTTA GTGGTAATTCTAGAGCTAATACT ATGTATTAGCTCTAGAATTACCAC CTGCCCTATCAACTTTCGA TCGAAAGTTGATAGGGCAG GTCTAGTATAATCGTTATACAGG GGAGTATGGTCGCAAGGCTG CAGCCTTGCGACCATACTCC GTGAGTTTCCCCGTGTTGAG TTGCTACATGGATAACTGTG CTGTTAAGACTACAACGG AGAGTGTTCAAAGCAGGC CTCACTAAGCCATTCAATCGG TCTAGAGCTAATACATGC(T/C)G GGCCCTCAAATCTAAGGATT 65 65 {62} 55 {72} {72} 59 53 53 55 60 74 61 50 57 64 58 57 57 49 52 53 57 45 51 49 53 53

  4. CD Genomics stands ready to provide reliable 18S characterization services, including 16S/18S/ITS amplicon sequencing by NGS and full-length 16S/18S/ITS sequencing by PacBio SMRT technology. Please contact our scientists for more detailed information. References: 1. Berkeley university of California (https://nature.berkeley.edu/brunslab/tour/primers.html#18s) 2. Buse H Y, Lu J, Lu X, et al. Microbial diversities (16S and 18S rRNA gene pyrosequencing) and environmental pathogens within drinking water biofilms grown on the common premise plumbing materials unplasticized polyvinylchloride and copper. FEMS microbiology ecology, 2014, 88(2): 280-295. 3. Wu S, Xiong J, Yu Y. Taxonomic resolutions based on 18S rRNA genes: a case study of subclass copepoda. PloS one, 2015, 10(6): e0131498. Tagged with: 18s rna 18s rna sequencing Fungal Diversity Analysis Fungal sequencing metagenomic analysis Primers for 18S rRNA

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