Callus Browning Research Articles

Unveiling the Molecular Mechanisms of Browning in Camellia hainanica Callus through Transcriptomic and Metabolomic Analysis.

Camellia hainanica is one of the camellia plants distributed in tropical regions, and its regeneration system and genetic transformation are affected by callus browning. However, the underlying mechanism of Camellia hainanica callus browning formation remains largely unknown. To investigate the metabolic basis and molecular mechanism of the callus browning of Camellia hainanica, histological staining, high-throughput metabolomics, and transcriptomic assays were performed on calli with different browning degrees (T1, T2, and T3). The results of histological staining revealed that the brown callus cells had obvious lignification and accumulation of polyphenols. Widely targeted metabolomics revealed 1190 differentially accumulated metabolites (DAMs), with 53 DAMs annotated as phenylpropanoids and flavonoids. Comparative transcriptomics revealed differentially expressed genes (DEGs) of the T2 vs. T1 associated with the biosynthesis and regulation of flavonoids and transcription factors in Camellia hainanica. Among them, forty-four enzyme genes associated with flavonoid biosynthesis were identified, including phenylalaninase (PAL), 4-coumaroyl CoA ligase (4CL), naringenin via flavanone 3-hydroxylase (F3H), flavonol synthase (FLS), Chalcone synthase (CHS), Chalcone isomerase (CHI), hydroxycinnamoyl-CoA shikimate transferase (HCT), Dihydroflavonol reductase (DFR), anthocyanin reductase (LAR), anthocyanin synthetase (ANS), and anthocyanin reductase (ANR). Related transcription factors R2R3-MYB, basic helix-loop-helix (bHLH), and WRKY genes also presented different expression patterns in T2 vs. T1. These results indicate that the browning of calli in Camellia hainanica is regulated at both the transcriptional and metabolic levels. The oxidation of flavonoids and the regulation of related structural genes and transcription factors are crucial decisive factors. This study preliminarily revealed the molecular mechanism of the browning of the callus of Camellia hainanensis, and the results can provide a reference for the anti-browning culture of Camellia hainanica callus.

Identification of QTLs responsible for culturability, and fine-mapping of QTL qCBT9 related to callus browning derived from Dongxiang common wild rice (Oryza rufipogon Griff.).

The online version contains supplementary material available at 10.1007/s11032-024-01470-z.

Genetic and Transcriptome Analyses of Callus Browning in Chaling Common Wild Rice (Oryza rufipogon Griff.).

Callus browning during tissue culture of indica rice is genotype dependent, thus limiting the application of genetic transformation for editing-assisted breeding and elucidation of gene function. Here, using 124 introgression lines (HCLs) derived from a cross between the indica rice 9311 and Chaling common wild rice and 2059 SNPs for single-point and interval analysis, we identified two major QTLs, qCBT7 on chromosome 7 and qCBT10 on chromosome 10, related to callus browning, explaining 8-13% of callus browning. Moreover, we performed RNA-seq of two introgression lines with low callus browning, HCL183 and HCL232, with Oryza. rufipogon introgression fragments on chromosomes 10 and 7, respectively. Three candidate genes (Os07g0620700, Os10g0361000, and Os10g0456800) with upregulation were identified by combining interval mapping and weighted gene coexpression network analysis using the DEGs. The qRT-PCR results of the three candidate genes were consistent with those of RNA-seq. The differentiation of indica and japonica subspecies Oryza. sativa and Oryza. rufipogon suggests that these candidate genes are possibly unique in Oryza. rufipogon. GO analyses of hub genes revealed that callus browning may be mainly associated with ethylene and hormone signaling pathways. The results lay a foundation for future cloning of qCBT7 or qCBT10 and will improve genetic transformation efficiency in rice.

Metabolite profiling and screening of callus browning-related genes in lotus (Nelumbo nucifera).

Callus browning is a major drawback to lotus callus proliferation and regeneration. However, the underlying mechanism of its formation remains largely unknown. Herein, we aimed to explore the metabolic and molecular basis of lotus callus browning by combining histological staining, high-throughput metabolomics, and transcriptomic assays for lotus callus at three browning stages. Histological stained brown callus cross sections displayed severe cell death symptoms, accompanied by an obvious accumulation of polyphenols and lignified materials. Widely targeted metabolomics revealed extensively decreased accumulation of most detected flavonoids and benzylisoquinoline alkaloids (BIAs), as well as a few phenolic acids, amino acids and their derivatives in callus with browning symptoms. Conversely, the contents of most detected tannins were significantly increased. Subsequent comparative transcriptomics identified a set of differentially expressed genes (DEGs) associated with the biosynthesis and regulation of flavonoids and BIAs in lotus. Notably, callus browning was coupled with significantly up-regulated expression of two polyphenol oxidase (PPO) and 17 peroxidase (POD) encoding genes, while the expression of ethylene associated genes remained at marginal levels. These results suggest that lotus callus browning is primarily controlled at the level of metabolism, wherein the oxidation of flavonoids and BIAs is crucially decisive.

Investigating the relationships between callus browning in Isatis indigotica Fortune, total phenol content, and PPO and POD activities

Investigating the relationships between callus browning in Isatis indigotica Fortune, total phenol content, and PPO and POD activities

A laccase gene (LcLac) was involved in polyphenol metabolism and tissue browning of litchi callus

Postharvest pericarp browning restricts the shelf life of litchi fruit and was considered as the main bottleneck of litchi industry. Browning is mainly due to the enzymatic catalysis of phenols to form brown substances in the litchi pericarp. (-)-Epicatechin, (-)-gallocatechin, and procyanidin B2 are the main substrates for enzyme catalysis. Previous works have reported that laccase activity was correlated with pericarp browning, and laccase polymerizes substrates in vitro, but the direct evidence was absent. In this research, the sequence of laccase gene LcLac was analyzed, whose expression was correlated with browning index of litchi pericarp at postharvest. Overexpression of LcLac in callus led to higher gene expression and enzymatic activity, and a lower polyphenol content, which contributed to browning of callus. Contrarily, the LcLac gene blocked by CRISPR/Cas9 resulted in an opposite trend of polyphenol contents. LcLac overexpression promoted callus browning, and the gene target block delayed the browning phenomenon. The results clearly indicate that LcLac can be polymerized with polyphenols and is involved in callus browning. LcLac may also participate in polyphenol synthesis and maintain the polyphenol balance in vacuoles.

Production of Secondary Metabolites from Cell Cultures of Sageretia thea (Osbeck) M.C. Johnst. Using Balloon-Type Bubble Bioreactors.

Sageretia thea is used in the preparation of herbal medicine in China and Korea; this plant is rich in various bioactive compounds, including phenolics and flavonoids. The objective of the current study was to enhance the production of phenolic compounds in plant cell suspension cultures of Sageretia thea. Optimum callus was induced from cotyledon explants on MS medium containing 2,4-dichlorophenoxyacetic acid (2,4-D; 0.5 mg L-1), naphthalene acetic acid (NAA, 0.5 mg L-1), kinetin (KN; 0.1 mg L-1) and sucrose (30 g L-1). Browning of callus was successfully avoided by using 200 mg L-1 ascorbic acid in the callus cultures. The elicitor effect of methyl jasmonate (MeJA), salicylic acid (SA), and sodium nitroprusside (SNP) was studied in cell suspension cultures, and the addition of 200 µM MeJA was found suitable for elicitation of phenolic accumulation in the cultured cells. Phenolic and flavonoid content and antioxidant activity were determined using 2,2 Diphenyl 1 picrylhydrazyl (DPPH), 2,2'-azino-bis (3-ethybenzothiazoline-6-sulphonic acid (ABTS), ferric reducing antioxidant power (FRAP) assays and results showed that cell cultures possessed highest phenolic and flavonoid content as well as highest DPPH, ABTS, and FRAP activities. Cell suspension cultures were established using 5 L capacity balloon-type bubble bioreactors using 2 L of MS medium 30 g L-1 sucrose and 0.5 mg L-1 2,4-D, 0.5 mg L-1 NAA, and 0.1 mg L-1 KN. The optimum yield of 230.81 g of fresh biomass and 16.48 g of dry biomass was evident after four weeks of cultures. High-pressure liquid chromatography (HPLC) analysis showed the cell biomass produced in bioreactors possessed higher concentrations of catechin hydrate, chlorogenic acid, naringenin, and other phenolic compounds.

Mapping quantitative trait loci associated with callus browning in Dongxiang common wild rice (Oryza rufipogon Griff.).

Genetic transformation of indica rice (Oryza sativa ssp. indica) is limited by callus browning, which results in poor in vitro tissue culturability. Elucidating the genes in common wild rice (Oryza rufipogon Griff.) that control callus browning is fundamental for improving the tissue culturability of indica rice varieties. We used a population of 129 O. rufipogon (Dongxiang common wild rice; DXCWR) introgression lines in the elite cultivar GC2 (Oryza sativa ssp. indica) background and 159 simple sequence repeat (SSR) markers to identify quantitative trait loci (QTLs) associated with callus browning. We evaluated callus browning based on the indices of callus browning rate (CBR), callus browning index (CBI), and standard callus browning index (SCBI). We detected 30 QTLs associated with callus browning across all lines, mapping to chromosomes 1, 2, 3, 4, 8, 9, and 12. These genomic regions were repeatedly associated with differences in CBR, CBI, and SCBI. The alleles from DXCWR showed additive effects in reducing callus browning. We identified new QTLs near the markers RM247 and RM7003 on chromosome 12, indicating that these QTLs are unique to DXCWR. Furthermore, we identified six introgression lines with significantly lower callus browning. These lines will be useful germplasms for genetic transformation and fine-mapping of the culturability trait.

Enhanced production of pinosylvin stilbene with aging of Pinus strobus callus and nematicidal activity of callus extracts against pinewood nematodes

Pinosylvin stilbenes are phenolic compounds mainly occurring in the Pinaceae family. We previously reported that the accumulation of two pinosylvin stilbene compounds, dihydropinosylvin methyl ether (DPME) and pinosylvin monomethyl ether (PME), in Pinus strobus trees was highly enhanced by infection with pine wood nematodes (PWNs: Bursaphelenchus xylophilus), and these two compounds showed strong nematicidal activity against PWNs. In this work, we established a system of pinosylvin stilbene (DPME and PME) production via the in vitro culture of P. strobus calli, and we examined the nematicidal activity of callus extracts. Calli were induced from the culture of mature zygotic embryos of P. strobus. Optimized growth of calli was obtained in 1/2 Litvay medium with 1.0 mg/L 2,4-D and 0.5 mg/L BA. DPME and PME accumulation did not occur in nonaged (one-month-old) calli but increased greatly with prolonged callus culture. The concentrations of DPME and PME in three-month-old dark-brown calli were 6.4 mg/g DW and 0.28 mg/g DW, respectively. The effect of methyl jasmonate treatment on the accumulation of DPME and PME was evaluated in cell suspension culture of P. strobus. However, the treatment appeared to show slight increase of DPME accumulation compared to callus browning. A test solution prepared from crude ethanol extracts from aged calli (three months old) containing 120 µg/ml DPME and 5.16 µg/ml PME treated with PWNs resulted in 100% immobilization of the adult PWNs and 66.7% immobilization of the juvenile PWNs within 24 h. However, nonaged callus extracts did not show any nematicidal activity against juvenile PWNs and showed less than 20% nematicidal activity against adult PWNs. These results indicate that pinosylvin stilbenes can be effectively produced by prolonged culture of P. strobus calli, can be isolated using simple ethanolic extraction, and are applicable as beneficial eco-friendly compounds with nematicidal activity against PWNs.

BREVIPEDICELLUS and ERECTA control the expression of AtPRX17 to prevent Arabidopsis callus browning.

Efficient in vitro callus generation is required for tissue culture propagation, a process that allows for plant regeneration and transgenic breeding for desired phenotypes. Identifying genes and regulatory elements that prevent impaired callus growth and callus browning is essential for the development of in vitro callus systems. Here, we show that the BREVIPEDICELLUS and ERECTA pathways in Arabidopsis calli converge to prevent callus browning, and positively regulate the expression of the isoperoxidase gene AtPRX17 in rapidly growing calli. Loss-of-function mutations in both BREVIPEDICELLUS and ERECTA resulted in markedly increased callus browning. Transgenic lines expressing 35S pro::AtPRX17 in the bp-5 er105 double mutant background fully rescued this phenotypic abnormality. Using in vivo (chromatin immunoprecipitation-PCR and transient expression) and in vitro (electrophoretic mobility shift assays) assays, we observed that the BREVIPEDICELLUS protein binds directly to the upstream sequence of AtPRX17 to promote its transcription during callus growth. ERECTA is a ubiquitous factor required for cell proliferation and growth. We show that ERECTA positively regulates the expression of the transcription factor WRKY6, which directly binds to a separate site on the AtPRX17 promoter, further increasing its expression. Our data reveal an important molecular mechanism involved in the regulation of peroxidase isozyme expression to reduce Arabidopsis callus browning.

Formamide deionized accelerates the somatic embryogenesis of Cunninghamia lanceolata

Aim of the study: To improve the efficiency of the somatic embryogenesis (SE) in Cunninghamia lanceolata. Area of the study: The study was conducted at Nanjing Forestry University (Nanjing, China). Material and methods: Immature cones of C. lanceolata, genotype 01A1 which was planted in Yangkou State-owned Forest Farm (Fujian, China), were used to induced callus. These calli were used to induce SE, concentration gradients of 0 g/L, 0.01134 g/L, 0.1134 g/L, 1.1134 g/L and 11.34 g/L of FD was added, to explore the optimal concentration for promoting SE of C. lanceolata. Main results: Low concentration of FD promoted the maturation of somatic embryos, while high concentration of FD lead to browning of embryogenic callus. The seedling rate and rooting number of seedlings induced by different concentrations of FD were significantly different. Research highlights: This study may aid in the rapid maturation of C. lanceolata somatic embryos and is useful for accelerated C. lanceolata breeding. Keywords: C. lanceolata; Formamide Deionized; Somatic embryogenesis; Seedling rate. Abbreviations used: FD (Formamide Deionized), FD0 (the concentration of 0 g/L FD), FD0.01134 (the concentration of 0.01134 g/L FD), FD0.1134 (the concentration of 0.1134 g/L FD), FD1.134 (the concentration of 1.134 g/L FD), FD11.34 (the concentration of 11.34 g/L FD).

Effect of Jasmonic Acid on Stimulating The Growth and Development of Date Palm Callus (Phoenix Dactylifera L.) Cultivar Shukar in Vitro Under Salt Stress Conditions

The study was conducted in the tissue culture laboratory at the Date Palm Research Center, University of Basrah. Using the date palm callus, Shukar cultivar, obtained from culturing the apical and axillary buds in sterile artificial nutrient media, to determine the effect of adding (0, 25, 50, 75) μM concentrations of Jasmonic acid (JA) to the MS media supplemented with several concentrations of NaCl (0, 50, 100, 200) mM were added, on the growth response and morphological development of callus after four months of cultivation in the propagation media. Results showed a significant decrease in growth characteristics in response to an increase in salt level, without the degree of callus browning that’s increased with the increase in the level of sodium chloride, especially at the two levels (100, 200) mM. Results showed also that the addition of Jasmonic acid to the saline treatments greatly enhanced the growth parameters of culture callus, as the rate of biomass, water content, relative growth rate, and the number of somatic embryos generated. as well as the degree of browning decreased significantly according to the levels of sodium chloride and the concentrations of added Jasmonic acid. The study demonstrated the effective role of exogenous addition of Jasmonic acid in vitro culture systems in enhancing stress tolerance of cells and tissues cultured in saline media.

Volatile organic compound and endogenous phytohormone characteristics during callus browning in Aquilaria sinensis

Callus browning, a common feature in plant cell cultures, has hardly been explored in Aquilaria sinensis. Here, the volatile organic compounds (VOCs) were characterized during the browning of A. sinensis cells using Gas Chromatography-Mass Spectrometry (GC–MS), and total sesquiterpene accumulation was significantly induced and peaked during A. sinensis callus browning, after 60 days of callus growth. Additionally, 14 types of sesquiterpenes were detected in the browning A. sinensis callus, with greater sesquiterpene diversity than in the ones treated with methyl jasmonate and salicylic acid (SA) in a previous study. To further evaluate the mechanism of regulation of agarwood sesquiterpenes during cell browning in A. sinensis, changes on phytohormone concentrations were explored using ultra-performance liquid chromatography-electrospray tandem mass spectrometry system (UPLC-ESI-MS/MS). Both endogenous jasmonic acid and SA, key plant defense hormones, increased during A. sinensis callus browning. 1-aminocyclopropanecarboxylic acid, an ethylene (ET) precursor, decreased significantly after A. sinensis callus browning. Conversely, two primary growth hormones, auxin and cytokinin, exhibited opposite trends, suggesting that they also play roles in agarwood sesquiterpene accumulation. Our results potentially offer novel insights that could facilitate the production valuable phytochemical compounds such as agarwood sesquiterpenes, using plant cell cultures treated with appropriate phytohormone combinations.

Effects of Light on Callus Multiplication of Actinidia Arguta

In order to screen out the best culture conditions for the growth of callus of Actinidia arguta, leaf callus and stem callus were used to explore the effect of different light intensity on callus multiplication. The results showed that, light could promote the synthesis of anthocyanins in callus, and low light intensity could keep the callus to maintain a healthy and physiological state with green color and compact structure. In addition, there was a negative correlation between light intensity and callus multiplication, and the stronger the light intensity, the more severe the callus browning. The optimal culture condition for leaf callus multiplication was dark environment, the optimal culture condition for stem callus multiplication was dark environment or 720 Lx light intensity.

An improved method for transformation of Actinidia arguta utilized to demonstrate a central role for MYB110 in regulating anthocyanin accumulation in kiwiberry

Actinidia arguta and related species produce small edible kiwifruit (kiwiberry) with attractive consumer features and high nutritional value. As a recently developed crop, kiwiberry can be improved further by classical breeding or genetic manipulation. The currently available Agrobacterium-mediated transformation protocol is only applicable to a limited number of kiwiberry genotypes such as the female cultivar ‘Hortgem Tahi’. We developed protocols for regeneration and Agrobacterium-mediated transformation of two A. arguta genotypes, female AA06-01 and male AA05-06, also applicable to ‘Hortgem Tahi’. Altered composition of basal salts in the callus-induction media was sufficient to prevent callus browning in AA06-01 and ‘Hortgem Tahi’, but not in AA05-06. Altering the hormone composition to avoid a prolonged callus stage was successfully used to prevent browning and induce shoot development in all three genotypes. Using our improved protocols, Agrobacterium-mediated transformation of a binary vector carrying the neomycin phosphotransferase II (nptII) expression cassette gave rise to kanamycin-resistant plants of both AA06-01 and AA05-06 and the presence of the nptII transgene was confirmed by genomic PCR. The improved protocols were also used to ectopically overexpress the anthocyanin-related transcription factor AcMYB110 in ‘Hortgem Tahi’ giving rise to purple callus with elevated anthocyanin accumulation. AcMYB110 expression and increased levels of anthocyanins were detected in mature leaves of established transgenic plants compared to controls, clearly demonstrating the important role for MYB110 in regulation of anthocyanin accumulation in kiwiberry. The protocols developed during this study provide tools for further functional analyses and genetic manipulation of kiwiberry genotypes. Using a newly developed transformation method we were able to generate transgenic lines of three Actinidia arguta genotypes and demonstrate an important role for MYB110 in anthocyanin accumulation in kiwiberry.

Effects of sodium nitroprusside and growth regulators on callus, multiple shoot induction and tissue browning in commercially important Valeriana jatamansi Jones

The effects of nitric oxide (NO) donor sodium nitroprusside (SNP) and various growth regulators on callus induction and shoot organogenesis in the commercially and pharmaceutically important medicinal plant Valeriana jatamansi is presented. Four different growth regulators at different concentrations were tested for their callusing efficiency followed by supplementation of SNP to the best responding growth regulator. Maximum callusing frequency (91.18%) was observed with the supplementation of 1.5 mg/L 1-Naphthaleneacetic acid + 15 μM SNP. 10% Coconut water (CW) was found to be the best growth regulator for the induction of shoots. The multiple shoot formation was also found to be enhanced with the supplementation of SNP. Maximum response for in vitro shoot multiplication was observed when the medium was supplemented with 10% CW + 15 μM SNP (89.32%) as compared to shoots induced in medium supplemented with 1.5 mg/L Benzyl aminopurine + 15 μM SNP (79.46%) respectively. No separate root induction medium was required as CW and CW + SNP supplementation was found to be sufficient enough to produce profuse rooting of the in vitro regenerated plants. Results revealed that SNP enhanced the callus and shoot induction frequencies. Supplementation of SNP also significantly reduced callus browing, allowed the tissues to recover and regenerate. This proposed in vitro culture system will be an effective mass propagation strategy for both commercial utilization and conservation of this highly valued plant species. First report using SNP and growth hormone supplementation for enhanced propagation in Valeriana jatamansi presented. Supplementation of SNP enhanced the callus, multiple shoot induction and reduced callus browning in the in vitro cultures of V. jatamansi.

A new approach to prevent hazelnut callus browning by modification of sub-culture

Tissue culture of hazelnut (Corylus avellana L.) represented the promising strategy for production of its valuable compound, paclitaxel, but one of the most important problems is initial callus browning in its callus culture. To obtain healthy callus cells of C. avellana, three different culture media, Murashige and Skoog (MS), Nas and Read (NRM), and Driver and Koniuki (DKW), in combination with 500 mg dm-3 citric acid, 500 mg dm-3 acetic acid, and 500 mg dm-3 polyvinylpyrrolidone (PVP) were applied. These cultures were passed two sub-cultures in the same media. Also, a novel modified sub-culture system was designed and compared to routine techniques. The sub-culture system was changed as following: the induced calli were transferred to a liquid media with the same composition and after developing the cell suspension, the cells were immobilized on a solid medium. In the first experiment, it was demonstrated that the medium compositions had an effect on the growth rate and callus browning reduction although they could not eliminate the browning. The results have shown that the highest growth indices were related to NRM with 500 mg dm-3 acetic acid + 500 mg dm-3 citric acid and to NRM with 500 mg dm-3 PVP in the first culture. These media had the least amount of browning (9.15 %). In the two next sub-cultures, NRM without additives had the maximum growth rate and the lowest browning was observed on the same media as in the first subculture (13.3 %). The technique of modified sub-cultures increases growth rate ten times in comparison with routine cultivation method.

Callus induction and browning suppression in tree peony Paeonia ostii ‘Fengdan’

Callus induction is an important stage in micropropagation. In this study, embryos, cotyledons, and hypocotyls of tree peony (Paeonia ostii ‘Fengdan’) were used as explants to induce callus formation. Callus induction was largely influenced by the medium, plant growth regulator, and explant. All combinations of the medium and plant growth regulator were conducive to callus induction from zygotic embryos, where the ratio of explants with callus induction was very high in all of the combinations. The greatest ratio of explants with callus induction from the cotyledon explants was found on Woody Plant Medium (WPM) or Murashige and Skoog (MS) medium supplemented with both 0.5 mg L−1 thidiazuron (TDZ) and either 0.5 mg L−1 2,4-dichlorophenoxyacetic acid (2,4-D) or 0.5 mg L−1 1-naphthaleneacetic acid (NAA), and on the WPM containing 0.5 mg L−1 NAA and 0.5 mg L−1 6-benzylaminopurine (BA). The ratio of explants with callus induced from the hypocotyl explants was the greatest on either MS medium or WPM supplemented with both 0.5 mg L−1 2,4-D and 0.5 mg L−1 TDZ. In contrast, a combination of 0.5 mg L−1 2,4-D and 0.5 mg L−1 TDZ, or 0.5 mg L−1 NAA and 0.5 mg L−1 TDZ was highly likely to cause a browning problem during culture. For browning suppression, calcium chloride (CaCl2), polyvinyl pyrrolidone (PVP), gallic acid, and caffeic acid were much more effective than other reagents. The efficacy on browning suppression was concentration-dependent, and the best results were obtained at a concentration of 4.0 mg L−1 CaCl2, 1.0–2.0 g L−1 PVP, 0.5–1.0 mg L−1 gallic acid, and 0.5–1.0 mg L−1 caffeic acid, respectively. In summary, callus formation was successfully induced from the zygotic embryo, cotyledon, and hypocotyl explants, and callus browning was effectively suppressed by caffeic acid (0.5–1.0 mg L−1), gallic acid (1.0 mg L−1), CaCl2 (4.0 mg L−1), and PVP (1.0 g L−1).

A common wild rice-derived BOC1 allele reduces callus browning in indica rice transformation

Callus browning, a common trait derived from the indica rice cultivar (Oryza sativa L.), is a challenge to transformation regeneration. Here, we report the map-based cloning of BROWNING OF CALLUS1 (BOC1) using a population derived from crossing Teqing, an elite indica subspecies exhibiting callus browning, and Yuanjiang, a common wild rice accession (Oryza rufipogon Griff.) that is less susceptible to callus browning. We show that BOC1 encodes a SIMILAR TO RADICAL-INDUCED CELL DEATH ONE (SRO) protein. Callus browning can be reduced by appropriate upregulation of BOC1, which consequently improves the genetic transformation efficiency. The presence of a Tourist-like miniature inverted-repeat transposable element (Tourist MITE) specific to wild rice in the promoter of BOC1 increases the expression of BOC1 in callus. BOC1 may decrease cell senescence and death caused by oxidative stress. Our study provides a gene target for improving tissue culturability and genetic transformation.

Transcriptome sequencing and identification of key callus browning-related genes from petiole callus of tree peony (Paeonia suffruticosa cv. Kao) cultured on media with three browning inhibitors

Tree peony (Paeonia suffruticosa Andrews) has ornamental, oil, and medicinal values, and demand in the markets for uniform tree peony seedlings is increasing. Micropropagation could quickly propagate uniform seedlings. However, the heavy browning phenomenon hinders large-scale development of uniform tree peony seedlings. In this paper, we measured the total phenolic compounds content, and sequenced the transcriptomes of tree peony ‘Kao’ petiole calluses cultured on media with three browning antagonist treatments and fresh petioles to identify the key genes involved in callus browning. Polyvinylpyrrolidone (PVP) treatment can reduce production of phenolic compounds and promote callus regeneration. A total of 218,957 unigenes were obtained from fresh petiole and three kinds of browning petiole calluses by transcriptome sequencing. The average sequence length of unigenes was 446 bp with an N50 of 493 bp. Functional annotation analysis revealed that 43,428, 45,357, 31,194, 30,019, and 21,357 unigenes were annotated using the NCBI-NR database, Swiss-Prot, KOG, GO, and KEGG, respectively. In total, 33 differentially expressed genes (DEGs) were identified as potentially associated with callus browning. Among these DEGs, 12 genes were predicted to participate in phenolic compounds biosynthesis, three genes were predicted to be involved in phenolic compounds oxidation, and six genes were predicted to participate in callus regeneration. Moreover, six transcription factors were observed to be differentially expressed in the fresh petiole and three treated petioles in tree peony. This study comprehensively identifies browning-related gene resources and will possibly help in deciphering the molecular mechanisms of callus browning of tree peony in the future.