Comparative expression of miRNA genes and miRNA-based AFLP marker analysis in cultivated tetraploid cottons.

Department of Plant and Environmental Sciences, New Mexico State University, MSC 3Q, Las Cruces, NM 88003, USA.
Journal of Plant Physiology (Impact Factor: 2.77). 12/2011; 168(8):824-30. DOI: 10.1016/j.jplph.2010.10.006
Source: PubMed

ABSTRACT MicroRNAs (miRNAs) are a class of small non-coding RNAs that down-regulate gene expression in a sequence specific manner to control plant growth and development. The identification and characterization of miRNAs are critical steps in finding their target genes and elucidating their functions. The objective of the present study was to assess the genetic variation of miRNA genes through expression comparisons and miRNA-based AFLP marker analysis. Seven miRNAs were first selected for RT-PCR and four for quantitative RT-PCR analysis that showed considerably high or differential expression levels in early stages of boll development. Except for miR160a, differential gene expression of miR171, 390a, and 396a was detected in early developing bolls at one or more timepoints between two cultivated cotton cultivars, Pima Phy 76 (Gossypium barbadense) and Acala 1517-99 (Gossypium hirsutum). Our further work demonstrated that genetic diversity of miRNA genes can be assessed by miRNA-AFLP analysis using primers designed from 22 conserved miRNA genes in combination with AFLP primers. Homologous miRNA genes can be also identified and isolated for sequencing and confirmation using this homology-based genotyping approach. This strategy offers an alternative to isolating a full length of miRNA genes and their up-stream and down-stream sequences. The significance of the expression and sequence differences of miRNAs between cotton species or genotypes needs further studies.

  • [Show abstract] [Hide abstract]
    ABSTRACT: miRNA is an exogenous non-coding RNA 21-24nt in length, which plays a crucial role in almost all biological processes. In plants, miRNAs regulate organ development, phase change, signal transduction and response to different biotic and abiotic stresses at the post-transcriptional levels. Although there are many studies on plant miRNAs, no studies have been focused on the genotype dependence. Genotype-dependent traits may be controlled by the differential expression of certain miRNAs. To test this hypothesis, we investigated the expression profile patterns of 11 selected miRNAs in 5 different organs in 5 different cotton cultivars and their implication on plant development. Our results demonstrate that miRNAs have different expression patterns in different plant organs in different genotypes, which implicate their different traits, including early flowering. miR172 is a miRNA controlling floral development and phase change; our results show that miR172 has a higher expression level in the flower bud than in any other organ, our results also show that Baimian cultivars have a higher expression of miR172 than TM-1. This suggests that Baimian cultivars have an earlier transition from vegetable growth to reproductive growth, which is confirmed by our development data on floral branch development. Our result also shows that several miRNAs, including miR159 and miR162, were highly expressed in Baimian cultivars. The results obtained in this study would provide new insight for improving cotton using miRNA-based biotechnology.
    Gene 06/2014; · 2.20 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: In plants, epigenetic regulation is important in normal development and in modulating some agronomic traits. The potential contribution of DNA methylation mediated gene regulation to phenotypic diversity and development in cotton was investigated between cotton genotypes and various tissues. DNA methylation diversity, genetic diversity, and changes in methylation context were investigated using methylation-sensitive amplified polymorphism (MSAP) assays including a methylation insensitive enzyme (BsiSI), and the total DNA methylation level was measured by high-performance liquid chromatography (HPLC). DNA methylation diversity was greater than the genetic diversity in the selected cotton genotypes and significantly different levels of DNA methylation were identified between tissues, including fibre. The higher DNA methylation diversity (CHG methylation being more diverse than CG methylation) in cotton genotypes suggest epigenetic regulation may be important for cotton, and the change in DNA methylation between fibre and other tissues hints that some genes may be epigenetically regulated for fibre development. The novel approach using BsiSI allowed direct comparison between genetic and epigenetic diversity, and also measured CC methylation level that cannot be detected by conventional MSAP.
    PLoS ONE 01/2014; 9(1):e86049. · 3.53 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: MicroRNAs (miRNAs) are small, endogenously expressed, non-coding RNA molecules involved in gene transcription and expression that combine with specific mRNA site of target genes to inhibit protein synthesis or degrade mRNA. Since the first plant miRNA was reported in 2002, numerous new miRNAs and their targets have been discovered via high-throughput sequencing and computational approaches. However, the genetic variation of miRNA genes is poorly understood due to the lack of miRNA-specific DNA markers. To study the genetic variation and map miRNAs and their putative target genes in cotton, we designed specific primers based on pre-miRNAs and published putative target genes. A total of 83 pre-miRNA primers and 1,255 putative target gene primers were surveyed, and 9 pre-miRNA polymorphic loci were mapped on 7 of the 26 tetraploid cotton chromosomes. Furthermore, 156 polymorphic loci of the target genes were mapped on the cotton genome. To map more miRNA loci, miRNA-based SRAP (sequence-related amplified polymorphism) markers were used to map an additional 54 polymorphic loci on the cotton genome with the exception of Chr01, Chr22, and Chr24. Finally, a network between miRNAs and their targets was constructed. All pre-miRNAs and 98 putative target genes were selected for RT-PCR analysis, revealing unique expression patterns across different fiber development stages between the mapping parents. Our data provide an overview of miRNAs, their putative targets, and their network in cotton as well as comparative expression analyses between Gossypium hirsutum and G. barbadense. These data provide a foundation for understanding miRNA regulation during cotton fiber development.
    BMC Genomics 10/2013; 14(1):706. · 4.40 Impact Factor


Available from
Aug 16, 2014