Regeneration in the Segmented Annelid Capitella teleta

The segmented worms, or annelids, are a clade within the Lophotrochozoa, one of the three bilaterian superclades. Annelids have long been models for regeneration studies due to their impressive regenerative abilities. Furthermore, the group exhibits variation in adult regeneration abilities with some species able to replace anterior segments, posterior segments, both or neither. Successful regeneration includes regrowth of complex organ systems, including the centralized nervous system, gut, musculature, nephridia and gonads. Here, regenerative capabilities of the annelid Capitella teleta are reviewed. C. teleta exhibits robust posterior regeneration and benefits from having an available sequenced genome and functional genomic tools available to study the molecular and cellular control of the regeneration response. The highly stereotypic developmental program of C. teleta provides opportunities to study adult regeneration and generate robust comparisons between development and regeneration.

Screening of Suitable Plant Regeneration Protocols for Several Capsicum spp. through Direct Organogenesis

Peppers (Capsicum spp.) are recalcitrant to in vitro culture regeneration, making the application of in vitro-based breeding strategies difficult. We evaluated the impact of different combinations of auxins, cytokinins and micronutrients on the induction of direct organogenesis in cotyledon and hypocotyl explants of C. annuum, C. baccatum and C. chinense. We found variation in the regeneration response among species and type of explant. In this way, the average numbers of shoots per cotyledon and hypocotyl explant were, respectively, 1.44 and 0.28 for C. annuum, 4.17 and 3.20 for C. baccatum and 0.08 and 0.00 for C. chinense. Out of the six media, the best overall results were obtained with the medium Pep1, which contained 5 mg/L BAP (6-benzylaminopurine), 0.5 mg/L IAA (indole-3-acetic acid) and 0.47 mg/L CuSO4, followed by a subculture in the same medium supplemented with 10 mg/L AgNO3 (medium Pep1.2). The best result for the Pep1 + Pep1.2 medium was obtained for C. baccatum using cotyledon explants, with 8.87 shoots per explant. The explants grown in medium Pep1 + Pep1.2 were the ones with greener tissue, while overall the hypocotyl explants were greener than the cotyledon explants. Our results indicate that there is wide variation among Capsicum species in terms of regeneration. Our results suggest that the synergistic effect of copper and silver resulted in a higher regeneration rate of Capsicum explants. Explants with shoots were transferred to different media for elongation, rooting and acclimatization. Although acclimatized plantlets were obtained for C. baccatum and C. chinense, an improvement in these latter stages would be desirable for a high throughput regeneration pipeline. This work contributes to the improvement of Capsicum regeneration protocols using specific combinations of medium, explant and genotype, reaching the levels of efficiency required for genetic transformation and of gene editing technologies for other crops.

Agrobacterium-mediated Genetic Transformation of Rice var. BRRI Dhan 58

Agrobacterium mediated genetic transformation of BRRI Dhan 58 was conducted by using immature embryos following indirect regeneration. High percentage of callus induction at 96.5% was obtained when seeds of BRRI dhan 58 were cultured on modified MS medium supplemented with 2.5 mg/l 2, 4-D under dark condition. The maximum regeneration response was rerecorded when MS was supplemented with 3 mg/l BAP + 0.5 mg/l NAA and 1.0 mg/l Kn. Genetic transformation was performed using A. tumefaciens strain LBA4404 harboring pCAMBIA1301 plasmid carrying the marker genes for β-glucuronidase (GUS) and hygromycin resistance (hptII). Integration of the GUS gene into the genome of the rice plants was confirmed by PCR. The leaf segments of the PCR positive transformed plants showed the expression of GUS. The results of this study would be an effective tool for crop improvement and functional studies of gene on rice plant. Plant Tissue Cult. & Biotech. 31(1): 71-80, 2021 (June)

Pars Pro Toto: Every Single Cell Matters

Compared to other species, plants stand out by their unparalleled self-repair capacities. Being the loss of a single cell or an entire tissue, most plant species are able to efficiently repair the inflicted damage. Although this self-repair process is commonly referred to as “regeneration,” depending on the type of damage and organ being affected, subtle to dramatic differences in the modus operandi can be observed. Recent publications have focused on these different types of tissue damage and their associated response in initiating the regeneration process. Here, we review the regeneration response following loss of a single cell to a complete organ, emphasizing key molecular players and hormonal cues involved in the model species Arabidopsis thaliana. In addition, we highlight the agricultural applications and techniques that make use of these regenerative responses in different crop and tree species.

Conservation, Regeneration and Genetic Stability of Regenerants from Alginate-Encapsulated Shoot Explants of Gardenia jasminoides Ellis

The present study demonstrates the potential of the alginate encapsulation of shoot tips and nodal segments of Gardenia jasminoides Ellis, the short-term cold storage of artificial seeds and subsequent successful conversion to desirable, uniform and genetically stable plantlets. Shoot tips and first-node segments below them, derived from shoots of in vitro cultures, responded better than second-to-fourth-node segments on agar-solidified Murashige and Skoog (MS) nutrient medium and thus, they were used as explants for alginate encapsulation. Explant encapsulation in 2.5% sodium alginate in combination with 50 mM of calcium chloride resulted in the production of soft beads, while hardening in 100 mM of calcium chloride formed firm beads of uniform globular shape, suitable for handling. The addition of liquid MS nutrient medium in the sodium alginate solution doubled the subsequent germination response of the beads. The maintenance of alginate beads under light favored their germination response compared to maintenance in darkness. Encapsulated shoot tip explants of gardenia, which were stored at 4 °C for 4, 8 or 12 weeks, showed a gradual decline in their regeneration response (73.3, 68.9, 53.3%, respectively), whereas, non-encapsulated explants (naked), stored under the same time durations of cold conditions, exhibited a sharp decline in regeneration response up to entirely zeroing (48.9, 11.1, 0.0%, respectively). Shoots, derived from 12-week cold-stored encapsulated explants, were easily rooted in solid MS nutrient medium with the addition of 0.5 μM of Indole-3-acetic acid (IAA) and after transplantation of the rooted plantlets individually to pots containing a peat–perlite (3:1, v/v) substrate, they were successfully acclimatized in the greenhouse under the gradual reduction of 75 or 50% shading with survival rates of 95–100%. The genetic stability of the acclimatized plantlets was assessed and compared with the mother plant using inter simple sequence repeat (ISSR) markers. ISSR analysis confirmed that all regenerated plantlets were genetically identical to the mother plant. This procedure of artificial seed production could be useful for the short-term storage of germplasm and the production of genetically identical and stable plants as an alternative method of micropropagation in Gardenia jasminoides.

Confirmation of “pre-plasmolysis mediated ex-osmosis hypothesis” to obtain shoot bud morphogenesis in Catharanthus roseus

The antineoplastic herb, Catharanthus roseus is a classified high-value low-volume medicinal herb which is in global attention of scientific research for modulation of its monoterpenoid indole alkaloids (MIA) pathway through genetic engineering. These secondary metabolites are generally stored in specific types of structures/compartments due to their cytotoxic nature and designated roles in plant defense response. However, their presence can hinder the genetic engineering process used to develop transgenic plants through de novo morphogenesis and regeneration of plants from cultured cells/tissues and hence, it always remained a critical impediment in transgenic research in C. roseus. The pre-plasmolysis treatment of leaf explants can help to tackle the recalcitrant nature of leaf explant and can support the direct regeneration response by ex-osmosis that minimizes the concentration of alkaloids. Therefore, this study was performed to chase the effect of osmotic conditions on recalcitrant leaves of C. roseus engaged in vitro plant regeneration and hypothesis of alkaloids ex-osmosis is confirmed by HPLC analysis.

Aging impairs the essential contributions of non-glial progenitors to neurorepair in the dorsal telencephalon of the Killifish N. furzeri

SummaryThe aging central nervous system (CNS) of mammals displays progressive limited regenerative abilities. Recovery after loss of neurons is extremely restricted in the aged brain. Many research models fall short in recapitulating mammalian aging hallmarks or have an impractically long lifespan. We established a traumatic brain injury model in the African turquoise killifish (Nothobranchius furzeri), a regeneration-competent vertebrate model that evolved to naturally age extremely fast. Stab-wound injury of the aged killifish dorsal telencephalon unveils an impaired and incomplete regeneration response when compared to young individuals. Remarkably, killifish brain regeneration is mainly supported by atypical non-glial progenitors, yet their proliferation capacity appears declined with age. We identified a high inflammatory response and glial scarring to also underlie the hampered generation of new neurons in aged fish. These primary results will pave the way for further research to unravel the factor age in relation to neurorepair, and to improve therapeutic strategies to restore the injured and/or diseased aged mammalian CNS.HighlightsAging impairs neurorepair in the killifish pallium at multiple stages of the regeneration processAtypical non-glial progenitors support the production of new neurons in the naive and injured dorsal palliumThe impaired regeneration capacity of aged killifish is characterized by a reduced reactive proliferation of these progenitors followed by a decreased generation of newborn neurons that in addition, fail to reach the injury siteExcessive inflammation and glial scarring surface as potential brakes on brain repair in the aged killifish pallium

Explants selection for in vitro propagation of Pachyrhizus erosus L.

Pachyrhizus erosus tuber is rich in protein asides its agronomical value as a legume, but the seeds by which it is propagated have very low viability. This study established sterilization protocol and effect of various concentrations of auxins and cytokinins on callus production and shoot regeneration from explants of P. erosus. Explants and seeds were sterilized using sodiumhypochlorite (NaClO) solution (5, 10 and 15% v/v) for 5 and 10 mins. Nodal, stem and leaf explants from in vitro germinated P. erosus and tuber from field grown plant were sterilized and cultured on Murashige and Skoog (MS) medium (control) and MS combined with different concentrations of auxins (NAA and 2, 4-D) and cytokinin (BA and Kinetin) and the cultured explants were monitored in terms of degree of callus formation, morphology and colour of callus and also for shoot induction. The results showed that seeds of P. erosus sterilized with 10% NaClO solution for 10 mins and germinated in vitro is the best way of getting sterile nodal, stem and leaf explants for the in vitro propagation of the plant, while tuber explants could be sterilized with 15% NaClO for 10 minutes. Nodal explants inoculated in MS medium supplemented with 1.0 mg/L BA gave the highest shoot regeneration response, while stem explants inoculated on MS medium supplemented with 1.0 mg/L BA and 0.5 mg/L NAA also gave the highest amount of friable callus. The study concluded that in vitro germinated seeds were the best way of getting explant for P. erosus.