Abstract
Genome size is one of the fundamental cytogenetic features of a species, which is critical for the design and initiation of any genome sequencing projects and can provide essential insights in studying taxonomy, cytogenetics, phylogenesis, and evolutionary studies. However, this key cytogenetic information is almost lacking in the endemic species Reseda pentagyna and the locally rare species Reseda lutea in Saudi Arabia. Therefore, genome size was analyzed by propidium iodide PI flow cytometry and compared to k-mer analysis methods. The standard method for genome size measures (flow cytometry) estimated the genome size of R. lutea and R. pentagyna with nuclei isolation MB01 buffer were found to be 1.91 ± 0.02 and 2.09 ± 0.03 pg/2 °C, respectively, which corresponded approximately to a haploid genome size of 934 and 1.022 Mbp, respectively. For validation, K-mer analysis was performed on both species’ Illumina paired-end sequencing data from both species. Five k-mer analysis approaches were examined for biocomputational estimation of genome size: A general formula and four well-known programs (CovEST, Kmergenie, FindGSE, and GenomeScope). The parameter preferences had a significant impact on GenomeScope and Kmergenie estimates. While the general formula estimations did not differ considerably, with an average genome size of 867.7 and 896. Mbp. The differences across flow cytometry and biocomputational predictions may be due to the high repeat content, particularly long repetitive regions in both genomes, 71% and 57%, which interfered with k-mer analysis. GenomeScope allowed quantification of high heterozygosity levels (1.04 and 1.37%) of R. lutea and R. pentagyna genomes, respectively. Based on our observations, R. lutea may have a tetraploid genome or higher. Our results revealed fundamental cytogenetic information for R. lutea and R. pentagyna, which should be used in future taxonomic studies and whole-genome sequencing.
Highlights
The development of advanced genomic technologies, and the subsequent storm of data from next-generation sequencing (NGS), has been a great asset to genomic research
Seeds from adult plants of both the endemic species R. pentagyna a R. lutea were collected from Abha, Saudi Arabia, for in vitro plant prop tification was confirmed through morphological features co3uopf 1l8ed wit Flora of Saudi Arabia [28] and protologue [29], and a voucher specime SBSN00016) was deposited at the Seed Bank Herbarium, College of Sc RU. lnutieva eborsthiteyxp, eKriSmAen.taTllhyeusiinngtaflcotwsceyetodmsewtryearnedscuomrfpauctaet-iosntealrlyiluizsiendg twheikth-m0er.3% sod afpoprro2achtothr3oumghianc,otmhbeinnatiwonaosfhsheodrt-3reatdose4quteinmciensg wwitihthbiodinofourmbaleti-css ttoeorlisl.ized wate 2g. eMramteriinalaatneddMoenth2od%s agar inoculated on Murashige and Skoog (MS)
This was demonstrated in investigations with cane toad [72], vanilla [73], and Pacific oyster [49,74], where k-mer-based GenomeScope estimations of genome sizes were barely half of those derived by flow cytometry and far smaller than those achieved after genome assembly
Summary
The development of advanced genomic technologies, and the subsequent storm of data from next-generation sequencing (NGS), has been a great asset to genomic research. Variations significant enough to differentiate a population into distinct species may still be difficult to discern employing classic morphological or DNA sequence; such variations may become more obvious when genome size is investigated along with other proofs [7,8]. Flow cytometry is a low-cost, relatively reliable, and quick laboratory technique for estimating plant genome size. Researchers can utilize many available programs to estimate genome size using sequencing data as well as the popular equation, i.e., the quotient of the k-mers total number and the peak frequency distribution. R. lutea L. is a deep-rooted biennial or perennial herbaceous plant that can grow up to 80 cm high and well adapted to fallow fields, rocky slopes, and roadsides It is distributed and spread throughout many temperate zones of the world [27]. Seeds from adult plants of both the endemic species R. pentagyna a R. lutea were collected from Abha, Saudi Arabia, for in vitro plant prop tification was confirmed through morphological features co3uopf 1l8ed wit Flora of Saudi Arabia [28] and protologue [29], and a voucher specime SBSN00016) was deposited at the Seed Bank Herbarium, College of Sc RU. lnutieva eborsthiteyxp, eKriSmAen.taTllhyeusiinngtaflcotwsceyetodmsewtryearnedscuomrfpauctaet-iosntealrlyiluizsiendg twheikth-m0er.3% sod afpoprro2achtothr3oumghianc,otmhbeinnatiwonaosfhsheodrt-3reatdose4quteinmciensg wwitihthbiodinofourmbaleti-css ttoeorlisl.ized wate 2g. eMramteriinalaatneddMoenth2od%s agar inoculated on Murashige and Skoog (MS)
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