Cytokinin, auxin, and abscisic acid affects sucrose metabolism conduce to de novo shoot organogenesis in rice (Oryza sativa L.) callus

Abstract

BackgroundShoot regeneration frequency in rice callus is still low and highly diverse among rice cultivars. This study aimed to investigate the association of plant hormone signaling and sucrose uptake and metabolism in rice during callus induction and early shoot organogenesis. The immatured seeds of two rice cultivars, Ai-Nan-Tsao 39 (ANT39) and Tainan 11 (TN11) are used in this study.ResultsCallus formation is earlier, callus fresh weight is higher, but water content is significant lower in ANT39 than in TN11 while their explants are inoculated on callus induction medium (CIM). Besides, the regeneration frequency is prominently higher in ANT39 (~80%) compared to TN11 callus (0%). Levels of glucose, sucrose, and starch are all significant higher in ANT39 than in TN11 either at callus induction or early shoot organogenesis stage. Moreover, high expression levels of Cell wall-bound invertase 1, Sucrose transporter 1 (OsSUT1) and OsSUT2 are detected in ANT39 at the fourth-day in CIM but it cannot be detected in TN11 until the tenth-day. It suggested that ANT39 has higher callus growth rate and shoot regeneration ability may cause from higher activity of sucrose uptake and metabolism. As well, the expression levels of ORYZA SATIVA RESPONSE REGULATOR 1 (ORR1), PIN-formed 1 and Late embryogenesis-abundant 1, representing endogenous cytokinin, auxin and ABA signals, respectively, were also up-regulated in highly regenerable callus, ANT39, but only ORR1 was greatly enhanced in TN11 at the tenth-day in CIM.ConclusionThus, phytohormone signals may affect sucrose metabolism to trigger callus initiation and further de novo shoot regeneration in rice culture.Electronic supplementary materialThe online version of this article (doi:10.1186/1999-3110-54-5) contains supplementary material, which is available to authorized users.

Osmotic stress stimulates shoot organogenesis in callus of rice (Oryza sativa L.) via auxin signaling and carbohydrate metabolism regulation

“…However, the cross-talk among these factors is still less discussed, especially at the molecular level. In the past decade, we have endeavored to establish a highly efficient regeneration system and tried to clarify the possible mechanisms of totipotency in rice callus (Huang and Liu 1998;Huang et al , 2006Huang et al , 2012Lee and Huang 2013). Two cultural steps, embryogenic and/or organogenic callus induction and shoot regeneration, are necessary to enhance the regeneration system from rice explants.…”

“…The collected calli were dried in a ventilating oven at 80°C for 48 h to constant weight. Water content (%) determination and shoot organogenesis frequency (%) evaluation were performed according to our previous studies (Huang et al 2012;Lee and Huang 2013). The results were obtained from at least three independent experiments.…”

“…The Glucose Assay Kit (GAGO-20, Sigma, USA) was used for glucose content determination. All the preparation and determination procedures are done according to Lee and Huang (2013). Each sample was tested at least 3 times.…”

“…The NormFinder program was used (http://moma.dk/normfinder-software) to normalize the expression levels of all target genes. All the gene-specific primers information is described at previous study (Lee and Huang 2013). All the amplified sequences are single product and the sequences are corrected after commercial DNA sequencing service (data not shown).…”

“…Total RNA was isolated from collected samples by the TRIzol reagent method (Invitrogen, USA) and treated with TURBO DNA-free DNase (Ambion, TX, USA) to remove residual genomic DNA (Lee and Huang 2013). Firststranded cDNA was synthesized from 1 lg total RNA with use of an oligo-dT primer (ImPro-II Reverse Transcription System, Promega, USA).…”