An SpG-Cas9-based cytosine base editor expands the scope of genome editing in carrot plants.

KEY MESSAGE: SpG Cas9 significantly expands the genome editing scope in carrot with NGN PAM recognition.

GRF-GIF duo and GRF-GIF-BBM: novel transformation methodologies for enhancing regeneration efficiency of genome-edited recalcitrant crops.

MAIN CONCLUSION: This review describes the potential use of two novel transformation methodologies, GRF-GIF and GRF-GIF-BBM, for improving the regeneration efficiency of genome-edited recalcitrant plants.

Genome-Wide Analysis of SPL/miR156 Module and Its Expression Analysis in Vegetative and Reproductive Organs of Oil Palm (Elaeis guineensis).

The SPL (SQUAMOSA-promoter binding protein-like) gene family is one of the largest plant transcription factors and is known to be involved in the regulation of plant growth, development, and stress responses. The genome-wide analysis of SPL gene members in a diverse range of crops has been elucidated. However, none of the genome-wide studies on the SPL gene family have been carried out for oil palm, an important oil-yielding plant. In this research, a total of 24 EgSPL genes were identified via a genome-wide approach. Phylogenetic analysis revealed that most of the EgSPLs are closely related to the Arabidopsis and rice SPL gene members. EgSPL genes were mapped onto the only nine chromosomes of the oil palm genome. Motif analysis revealed conservation of the SBP domain and the occurrence of 1-10 motifs in EgSPL gene members. Gene duplication analysis demonstrated the tandem duplication of SPL members in the oil palm genome. Heatmap analysis indicated the significant expression of SPL genes in shoot and flower organs of oil palm plants. Among the identified EgSPL genes, a total 14 EgSPLs were shown to be targets of miR156. Real-time PCR analysis of 14 SPL genes showed that most of the EgSPL genes were more highly expressed in female and male inflorescences of oil palm plants than in vegetative tissues. Altogether, the present study revealed the significant role of EgSPL genes in inflorescence development.

Identification and expression analysis of histone modification gene (HM) family during somatic embryogenesis of oil palm.

BACKGROUND: Oil palm (Elaeis guineensis, Jacq.) is an important vegetable oil-yielding plant. Somatic embryogenesis is a promising method to produce large-scale elite clones to meet the demand for palm oil. The epigenetic mechanisms such as histone modifications have emerged as critical factors during somatic embryogenesis. These histone modifications are associated with the regulation of various genes controlling somatic embryogenesis. To date, none of the information is available on the histone modification gene (HM) family in oil palm. RESULTS: We reported the identification of 109 HM gene family members including 48 HMTs, 27 HDMs, 13 HATs, and 21 HDACs in the oil palm genome. Gene structural and motif analysis of EgHMs showed varied exon-intron organization and with conserved motifs among them. The identified 109 EgHMs were distributed unevenly across 16 chromosomes and displayed tandem duplication in oil palm genome. Furthermore, relative expression analysis showed the differential expressional pattern of 99 candidate EgHM genes at different stages (non-embryogenic, embryogenic, somatic embryo) of somatic embryogenesis process in oil palm, suggesting the EgHMs play vital roles in somatic embryogenesis. Our study laid a foundation to understand the regulatory roles of several EgHM genes during somatic embryogenesis. CONCLUSIONS: A total of 109 histone modification gene family members were identified in the oil palm genome via genome-wide analysis. The present study provides insightful information regarding HM gene's structure, their distribution, duplication in oil palm genome, and also their evolutionary relationship with other HM gene family members in Arabidopsis and rice. Finally, our study provided an essential role of oil palm HM genes during somatic embryogenesis process.

Transcriptome analysis of oil palm pistil during pollination and fertilization to unravel the role of phytohormone biosynthesis and signaling genes.

Phytohormones play an important role in the pollination and fertilization of crops, but the regulatory mechanisms of oil palm pollination and fertilization are unclear. The purpose of this study is to explore the hormonal changes of oil palm pistils during flowering. We used RNA sequencing to evaluate differentially expressed genes (DEGs) in oil palm pistils at the pollination and non-pollination stages. In this study, we found that the hormone contents of oil palm pistil changed drastically after pollination. The transcriptome of the oil palm pistil without pollination and at 2 h, 4 h, 12 h, 24 h, and 48 h after pollination was comprehensively analyzed, and a large number of differential genes and metabolic pathways were explored. Based on the transcriptome data, it could be recognized that the changes of indoleacetic acid (IAA), zeatin riboside (ZR), and abscisic acid (ABA) during pollination were consistent with the changes in the corresponding gene transcripts. Differentially expressed genes during pollination and fertilization of oil palm were mainly related to energy metabolism and hormone signal transduction. It provides new insights to elucidate the interaction and regulation mechanisms of plant hormones before and after oil palm pollination, providing a theoretical basis and reference for the research on sexual reproduction of oil palm.

Transcriptome analysis reveals key developmental and metabolic regulatory aspects of oil palm (Elaeis guineensis Jacq.) during zygotic embryo development.

BACKGROUND: Oil palm is the most efficient oil-producing crop in the world, and the yield of palm oil is associated with embryonic development. However, a comprehensive understanding of zygotic embryo development at the molecular level remains elusive. In order to address this issue, we report the transcriptomic analysis of zygotic embryo development in oil palm, specifically focusing on regulatory genes involved in important biological pathways. RESULTS: In this study, three cDNA libraries were prepared from embryos at S1 (early-stage), S2 (middle-stage), and S3 (late-stage). There were 16,367, 16,500, and 18,012 genes characterized at the S1, S2, and S3 stages of embryonic development, respectively. A total of 1522, 2698, and 142 genes were differentially expressed in S1 vs S2, S1 vs S3, and S2 vs S3, respectively. Using Gene Ontology (GO) term enrichment and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis to identify key genes and pathways. In the hormone signaling pathway, genes related to auxin antagonize the output of cytokinin which regulates the development of embryo meristem. The genes related to abscisic acid negatively regulating the synthesis of gibberellin were strongly up-regulated in the mid-late stage of embryonic development. The results were reported the early synthesis and mid-late degradation of sucrose, as well as the activation of the continuous degradation pathway of temporary starch, providing the nutrients needed for differentiation of the embryonic cell. Moreover, the transcripts of genes involved in fatty acid synthesis were also abundantly accumulated in the zygotic embryos. CONCLUSION: Taken together, our research provides a new perspective on the developmental and metabolic regulation of zygotic embryo development at the transcriptional level in oil palm.

The oil palm R2R3-MYB subfamily genes EgMYB111 and EgMYB157 improve multiple abiotic stress tolerance in transgenic Arabidopsis plants.

KEY MESSAGE: We found that overexpression of EgMYB111 and EgMYB157 genes positively regulate the abiotic stress tolerance. MYB family genes are well-known regulators in modulating the abiotic stress-responsive mechanisms in plants. However, lesser is known about the functional roles of oil palm MYB genes. Previously, we found that oil palm MYB genes such as EgMYB111 and EgMYB157 were significantly up-regulated under salinity, cold, and drought stress conditions. In this study, we over-expressed EgMYB111 and EgMYB157 genes separately in Arabidopsis plants. The transgenic Arabidopsis plants expressing EgMYB111 have shown improved tolerance to salinity, cold and drought stress conditions, whereas transgenic Arabidopsis plants expressing EgMYB157 dispalyed improved tolerance to cold and drought stress conditions only. Various biochemical analyses also revealed significant improvement of antioxidant enzyme activities, photosynthetic pigments, net photosynthetic rate, stomatal conductance, and intercellular CO2 concentration in transgenic plants compared to wild-type plants under cold, drought, and salinity stress conditions. Significant up-regulation of various known stress marker genes such as RD22, RD29A, RAB18, COR47, ABA1, ABI1, HAB1 was also noticed in EgMYB111 and EgMYB157 expressing transgenic plants compared to wild-type plants under cold, drought, and salinity stress conditions. Taken together, over-expression of EgMYB111 and/or EgMYB157 significantly improve abiotic tolerance in transgenic Arabidopsis plants, indicating that EgMYB111 and EgMYB157 are the potential candidates for developing abiotic stress-tolerant crops in near future.

The NAC-type transcription factor GmNAC20 improves cold, salinity tolerance, and lateral root formation in transgenic rice plants.

NAC-type transcription factors are crucial players in the abiotic stress responses of plants. Soybean NAC-type transcription factor GmNAC20 was transformed into rice genome via Agrobacterium method of transformation to improve abiotic stress tolerance. Integration and expression of GmNAC20 were verified by the DNA blot hybridization, immunoblotting, RT-PCR, and quantitative RT-PCR in T3 generation of transgenic rice plants. Significant expression of GmNAC20 was found in transgenic plants under salinity, cold, and IAA treatments. The transgenic rice plants expressing GmNAC20 displayed enhanced salinity and cold stress tolerance via upregulating the abiotic stress-responsive genes. Furthermore, T3 transgenic plants retained relative water content, chlorophyll content with enhanced accumulation of proline content than wild-type plants under salinity, and cold stress environments. The decrease in MDA content and electrolyte leakage with a significant increase in antioxidant enzyme activities were noticed in transgenic rice plants under either salinity or cold stress conditions, compared to wild-type plants. Overexpression of GmNAC20 in rice plants also induced the lateral root formation, associated with upregulation of auxin signaling-related genes. Taken together, our results indicated that GmNAC20 acts as a positive regulator for conferring salinity and cold tolerance in rice plants and appropriate candidate for improving salinity and cold stress in other important food crops.

Base editing in rice: current progress, advances, limitations, and future perspectives.

KEY MESSAGE: Base editing is one of the promising genome editing tools for generating single-nucleotide changes in rice genome. Rice (Oryza sativa L.) is an important staple food crop, feeding half of the population around the globe. Developing new rice varieties with desirable agronomic traits is necessary for sustaining global food security. The use of genome editing technologies for developing rice varieties is pre-requisite in the present scenario. Among the genome editing technologies developed for rice crop improvement, base editing technology has emerged as an efficient and reliable tool for precise genome editing in rice plants. Base editing technology utilizes either adenosine or cytidine base editor for precise editing at the target region. A base editor (adenosine or cytidine) is a fusion of catalytically inactive CRISPR/Cas9 domain and adenosine or cytidine deaminase domain. In this review, authors have discussed the different adenine and cytosine base editors developed so far for precise genome editing of rice via base editing technology. We address the current progress, advances, limitations, as well as future perspectives of the base editing technology for rice crop improvement.

Genome-Wide Identification and Characterization of AP2/ERF Transcription Factor Family Genes in Oil Palm under Abiotic Stress Conditions.

The AP2/ERF transcription factor family members play crucial roles in controlling plant growth and development, as well as responses to various abiotic stresses. Genome-wide identification and characterization of AP2/ERF genes has not yet been carried out in the oil palm genome. In the present work, we reported the occurrence of 172 EgAP2/ERFs (AP2, ERF, RAV & Soloist members) through genome-wide identification. Phylogenetic analysis was used to divide them into four groups, including: 34 AP2, 131 ERF, 5 RAV, and 2 Soloist gene family members. All 172 AP2/ERF members were unevenly distributed across 16 chromosomes of oil palm. Gene duplication analysis elucidated the tandem duplication of AP2/ERFs on chromosome blocks of the oil palm genome during evolution. Gene structure as well as conserved motif analysis demonstrated the conserved nature of intron/exon organization and motifs among the AP2/ERF genes. Several cis-regulatory elements-related to hormone, stress, and defense responses-were identified in the promoter regions of AP2/ERFs. Tissue-specific expression of 172 AP2/ERFs in five different tissues of oil palm was also revealed by heatmap analysis using the available transcriptome data. Finally, abiotic stress (salinity, cold & drought)-responsive AP2/ERFs in the oil palm genome were validated through qPCR analysis. Our study provided valuable information on oil palm AP2/ERF superfamily members and dissected their role in abiotic stress conditions.

Plastome engineering in vegetable crops: current status and future prospects.

Plastome (plastid genome) engineering has grown up and got smarter for the transgene expression. Plastid transformation has profound benefits over nuclear transformation, includes a higher level of transgene expression, integration via homologous recombination, transgene containment, lack of gene silencing, and position effect. Substantial and fruitful progress has been achieved in plastome engineering of vegetable crops through the use of improved regeneration/selection procedures, plastid transformation vectors with efficient promoters, and 3/, 5/regulatory sequences. Plastid transformation technology developed for vegetable crops being used as a platform for the production of industrially important proteins and some of the genes of agronomic importance has been stably integrated and expressed in plastome. Although great progress has been accomplished in the plastid transformation of vegetable crops, still it is restricted to few species because of the unavailability of whole plastome sequencing. In this review, the author focus on the technology, progress, and advancements in plastid transformation of vegetable plants such as lettuce, tomato, potato, cabbage, cauliflower, eggplant, carrot, soybean, and bitter melon are reviewed. The conclusions, future prospects, and expansion of plastid transformation technology to other vegetable crops for genetic improvement and production of edible vaccines are proposed.

Ectopic expression of nucleolar DEAD-Box RNA helicase OsTOGR1 confers improved heat stress tolerance in transgenic Chinese cabbage.

KEY MESSAGE: The DEAD-Box RNA helicase OsTOGR1 positively regulates heat stress tolerance in Chinese cabbage. Non-heading Chinese cabbage (Brassica rapa L. ssp. chinensis) is primarily cultivated vegetable crop in Asian countries. Heat stress is one of the major threats for its growth and yield. Numerous regulatory genes in various crops have shown to contribute thermotolerance. Among them, Thermotolerant growth required 1 (TOGR1) is an important DEAD-box RNA helicase. To examine whether its role is conserved in other crops, we constructed pCAMBIA1300-pHSP:OsTOGR1 expression vector driven by the rice small heat shock protein promoter (pHSP17.9) and successfully produced transgenic non-heading Chinese cabbage plants expressing OsTOGR1 gene via Agrobacterium-mediated vacuum infiltration transformation. In total, we generated three independent transgenic cabbage lines expressing TOGR1 gene. Expression and integration of TOGR1 was confirmed by PCR, RT-PCR and qPCR in T1 and T2 generations. The relative leaf electrical conductivity of transgenic seedlings was reduced subjected to high temperature (38 °C) compared to heat shock treatment (46 °C). In addition, hypocotyl length of transgenic seedlings increased compared to wild-type plants under high temperature and heat shock treatment. Furthermore, the transgenic plants exhibited higher chlorophyll content than wild-type plants under high temperature and heat shock treatment. The transgenic seeds displayed better germination under heat shock treatment. Tested heat stress-responsive genes were also up-regulated in the transgenic plants subjected to high temperature or heat shock treatment. To the best of our knowledge, this is the first report on describing the role of DAED-Box RNA helicases in improving heat stress tolerance of transgenic plants.

Expressing class I wheat NHX (TaNHX2) gene in eggplant (Solanum melongena L.) improves plant performance under saline condition.

Brinjal or eggplant (Solanum melongena L.) is an important solanaceous edible crop, and salt stress adversely affects its growth, development, and overall productivity. To cope with excess salinity, vacuolar Na+/H+ antiporters provide the best mechanism for ionic homeostasis in plants under salt stress. We generated transgenic eggplants by introducing wheat TaNHX2 gene that encodes a vacuolar Na+/H+ antiporter in to the eggplant genome via Agrobacterium-mediated transformation using pBin438 vector that harbors double35S:TaNHX2 to confer salinity tolerance. Polymerase chain reaction and southern hybridization confirmed the presence and integration of TaNHX2 gene in T1 transgenic plants. Southern positive transgenic eggplants showed varied levels of TaNHX2 transcripts as evident by RT-PCR and qRT-PCR. Stress-inducible expression of TaNHX2 significantly improved growth performance and Na+ and K+ contents from leaf and roots tissues of T2 transgenic eggplants under salt stress, compared to non-transformed plants. Furthermore, T2 transgenic eggplants displayed the stable leaf relative water content and chlorophyll content, proline accumulation, improved photosynthetic efficiency, transpiration rate, and stomatal conductivity than the non-transformed plants under salinity stress (200 mM NaCl). Data showed that the T2 transgenic lines revealed that reduction in MDA content, hydrogen peroxide, and oxygen radical production associated with the significant increase of antioxidant enzyme activity in transgenic eggplants than non-transformed plants under salt stress (200 mM NaCl). This study suggested that the TaNHX2 gene plays an important regulatory role in conferring salinity tolerance of transgenic eggplant and thus may serve as a useful candidate gene for improving salinity tolerance in other vegetable crops.

Improved salinity tolerance and growth performance in transgenic sunflower plants via ectopic expression of a wheat antiporter gene (TaNHX2).

Sunflower (Helianthus annuus. L) is one of the principal oil seed crops affected by the salinity stress, which limits the oil content and crop yield of sunflower plants. The acclimatization of plants to abiotic stresses such as salinity tolerance is mainly mediated by the vacuolar Na+/H+ antiporters (NHX) by tagging Na+ into vacuoles from the cytosol. We show here that the over-expression of wheat TaNHX2 gene in transgenic sunflower conferred improved salinity stress tolerance and growth performance. Transgenic sunflower plants were produced by infecting the embryonic axis ex-plants with Agrobacterium tumefaciens strain EHA105 containing a pBin438-TaNHX2 binary vector that carried a wheat antiporter (TaNHX2) gene under the control of a double CaMV 35S promoter with NPT II gene as a selectable marker. PCR analysis of T0 and T1 transgenic plants confirmed the integration of TaNHX2 in sunflower genome. Stable integration and expression of TaNHX2 in sunflower genome was further verified by Southern hybridization and semi-quantitative RT-PCR analyses. As compared to the non-transformed plants, TaNHX2 expressing transgenic plants showed better growth performance and accumulated higher Na+, K+ contents in leaves and roots under salt stress (200 mM NaCl). Transgenic sunflower plants displayed improved protection against cell damage exhibiting stable relative water content, chlorophyll content, increased proline accumulation and improved reactive oxygen species (ROS) scavenging because of higher activities of the antioxidant enzymes like superoxide dismutase and ascorbate peroxidase, along with decreased production of hydrogen peroxide, free oxygen radical and malondialdehyde (MDA) under salt stress (200 mM NaCl). Taken together, our findings suggest that TaNHX2 expression in sunflower plants contributed towards improving growth performance under sodium chloride stress.

Progress in Tissue Culture and Genetic Transformation of Oil Palm: An Overview.

Oil palm (Elaeis guineensis, Jacq.) is a prominent vegetable-oil-yielding crop. Cultivating high-yielding oil palm with improved traits is a pre-requisite to meet the increasing demands of palm oil consumption. However, tissue culture and biotechnological approaches can resolve these concerns. Over the past three decades, significant research has been carried out to develop tissue culture and genetic transformation protocols for oil palm. Somatic embryogenesis is an efficient platform for the micropropagation of oil palm on a large scale. In addition, various genetic transformation techniques, including microprojectile bombardment, Agrobacterium tumefaciens mediated, Polyethylene glycol mediated mediated, and DNA microinjection, have been developed by optimizing various parameters for the efficient genetic transformation of oil palm. This review mainly emphasizes the methods established for in vitro propagation and genetic transformation of oil palm. Finally, we propose the application of the genome editing tool CRISPR/Cas9 to improve the various traits in this oil yielding crop.

Combined transcriptome and metabolome analysis of sugar and fatty acid of aromatic coconut and non-aromatic coconut in China.

Sugar and fatty acid content are among the important factors that contribute to the intensity of flavor in aromatic coconut. Gaining a comprehensive understanding of the sugar and fatty acid metabolites in the flesh of aromatic coconuts, along with identifying the key synthetic genes, is of significant importance for improving the development of desirable character traits in these coconuts. However, the related conjoint analysis of metabolic targets and molecular synthesis mechanisms has not been carried out in aromatic coconut until now. UPLC-MS/MS combined with RNA-Seq were performed in aromatic coconut (AC) and non-aromatic coconut (NAC) meat at 7, 9 and 11 months. The results showed that D-fructose in AC coconut meat was 3.48, 2.56 and 3.45 fold higher than that in NAC coconut meat. Similarly, D-glucose in AC coconut meat was 2.48, 2.25 and 3.91 fold higher than that in NAC coconut meat. The NAC coconut meat showed a 1.22-fold rise in the content of lauric acid compared to the AC coconut meat when it reached 11 months of age. Myristic acid content in NAC coconut meat was 1.47, 1.44 and 1.13 fold higher than that in AC coconut meat. The palmitic acid content in NAC coconut meat was 1.62 and 1.34 fold higher than that in AC coconut meat. The genes SPS, GAE, GALE, GLCAK, UGE, UGDH, FBP, GMLS, PFK, GPI, RHM, ACC, FabF, FatA, FabG, and FabI exhibited a negative correlation with D-fructose (r = -0.81) and D-glucose (r = -0.99) contents, while showing a positive correlation (r = 0.85-0.96) with lauric acid and myristic acid. Furthermore, GALE, GLCAK, FBP, GMLS, and ACC displayed a positive correlation (r = 0.83-0.94) with palmitic acid content. The sugar/organic acid ratio exhibited a positive correlation with SPS, GAE, UGE, FabF, FabZ and FabI.

Genome-wide identification and expression analysis of bZIP transcription factors in oil palm (Elaeis guineensis Jacq.) under abiotic stress.

The bZIP transcription factors are well-known transcription regulators and play a key role in regulating various developmental, biological processes, and stress responses in plants. However, information on bZIP transcription factors is not yet available in oil palm, an important oil yielding crop. The present study identified the 97 bZIP transcription factor family members in oil palm genome via a genome-wide approach. Phylogenetic analysis clustered all EgbZIPs into 12 clusters with Arabidopsis and rice bZIPs. EgbZIP gene structure analysis showed a distinct variation in the intron-exon organization among all EgbZIPs. Conserved motif analysis demonstrated the occurrence of ten additional conserved motifs besides having a common bZIP domain. All the identified 97 EgbZIPs were unevenly distributed on 16 chromosomes and exhibited tandem duplication in oil palm genome. Our results aslo demonstrated that tissue-specific expression patterns of EgbZIPs based on the available transcriptome data of six different tissue of oil palm. Stress-responsive expression analysis showed that 11EgbZIP transcription factors were highly expressed under cold, salinity, drought stress conditions. Taken together, our findings will provide insightful information on bZIP transcription factors as one of the stress-responsive regulators in oil palm.

The auxin response factor (ARF) gene family in Oil palm (Elaeis guineensis Jacq.): Genome-wide identification and their expression profiling under abiotic stresses.

Auxin response factors (ARFs) play vital role in controlling growth and developmental processes of plants via regulating the auxin signaling pathways. However, the identification and functional roles of ARFs in oil palm plants remain elusive. Here, we identified a total of 23 ARF (EgARF) genes in oil palm through a genome-wide identification approach. The EgARF gene structure analysis revealed the presence of intron-rich ARF gene family in genome of oil palm. Further analysis demonstrated the uneven distribution of 23EgARFs on 16 chromosomes of oil palm. Phylogenetic analysis clustered all the EgARFs into four groups. Twenty-one EgARFs contained BDD, ARF, and CTD domains, whereas EgARF5 and EgARF7 lacked the CTD domain. The evolution of ARF genes in oil palm genome has been expanded by segmental duplication events. The cis-acting regulatory elements of EgARF gene family were predominantly associated with the stress and hormone responses. Expression profiling data demonstrated the constitutive and tissue-specific expression of EgARF genes in various tissues of oil palm. Real-time PCR analysis of 19 EgARF genes expression levels under cold, drought, and salt stress conditions proved their prominent role under abiotic stress responses. Altogether, our study provides a basis for studying the molecular and functional roles of ARF genes in oil palm.

Oil Palm Breeding in the Modern Era: Challenges and Opportunities.

Oil palm, a cross-pollinated crop with long generation time, poses a lot of challenges in achieving sustainable oil palm with high yield and quality. The African oil palm (Elaeis guineensis Jacq.) is the most productive and versatile oil-yielding crop in the world, producing more than any other oil-yielding crop. Despite recent challenges, such as stress tolerance, superior oil quality, disease tolerance, and the need for new market niches, there is a growing need to explore and develop new varieties with high yield potential and the genetic diversity required to maintain oil palm yield stability. Breeding is an indispensable part of producing high-quality planting materials to increase oil palm yield. Biotechnological technologies have transformed conventional plant breeding approaches by introducing novel genotypes for breeding. Innovative pre-breeding and breeding approaches, such as identifying candidate genes in wild or land races using genomics tools, can pave the way for genetic improvement in oil palm. In this review, we highlighted the modern breeding tools, including genomics, marker-assisted breeding, genetic engineering, and genome editing techniques in oil palm crops, and we explored certain concerns connected to the techniques and their applications in practical breeding.

Identification and function prediction of iron-deficiency-responsive microRNAs in citrus leaves.

Iron is a critical micronutrient for growth and development of plants and its deficiency limiting the crop productivity. MicroRNAs (miRNAs) play vital roles in adaptation of plants to various nutrient deficiencies. However, the role of miRNAs and their target genes related to Fe-deficiency is limited. In this study, we identified Fe-deficiency-responsive miRNAs from citrus. In Fe-deficiency conditions, about 50 and 31 miRNAs were up-regulated and down-regulated, respectively. The differently expressed miRNAs might play critical roles in contributing the Fe-deficiency tolerance in citrus plants. The miRNAs-mediated Fe-deficiency tolerance in citrus plants might related to the enhanced stress tolerance by decreased expression of miR172; regulation of S homeostasis by decreased expression of miR395; inhibition of plant growth by increased expression of miR319 and miR477; regulation of Cu homeostasis as well as activation of Cu/Zn superoxide dismutase activity due to decreased expression of miR398 and miR408 and regulation of lignin accumulation by decreased expression of miR397 and miR408. The identified miRNAs in present study laid a foundation to understand the Fe-deficiency adaptive mechanisms in citrus plants. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s13205-021-02669-z.