Comparative Analysis of Physical and Chemical Mutagenesis in Chrysanthemum cv. 'Candid': Assessing Genetic Variation and Breeding Potential.

In this study, we developed a mutagenesis protocol specifically designed for chrysanthemum cv. "Candid" in order to introduce genetic variation. By subjecting chrysanthemum shoots to different doses of physical and chemical mutagens, we successfully generated a total of 24 mutants, each with unique genetic compositions. We observed that the mortality rate was lowest when the shoots were exposed to 10 Gy gamma irradiation and 1.00% EMS. To assess the diversity and relatedness among the mutants, we employed RAPD and SSR markers. The combination of these markers allowed us to construct a dendrogram that effectively categorized the mutant population into distinct clusters based on the specific mutagen treatments. Interestingly, the mutants induced by 10 Gy gamma irradiation exhibited greater genetic diversity in terms of flower colors. On the other hand, mutants created with 1.00% EMS displayed a higher level of variation and yielded more viable mutants. To determine the optimal markers for studying genetic diversity, we analyzed the polymorphic information content (PIC) of different markers. Among the tested markers, OPA-07 (RAPD) and JH47 (SSR) showed the highest PIC values, indicating their effectiveness in capturing genetic variability within the mutant population. Conversely, the PIC values of OPD-07 and JH20 demonstrated the lowest among the markers tested. Our results revealed a percentage of polymorphism ranging from 81.81% to 100% for RAPD markers and 66.66% to 100% for SSR markers. These findings indicate that physical mutation induced by 10 Gy gamma irradiation can be clearly distinguished from chemical mutation induced by EMS at concentrations of 1% and 0.75% in chrysanthemum cv. "Candid.″ Overall, this study provides valuable insights into the genetic composition of the generated mutants and highlights their potential for enhancing chrysanthemum-breeding programs. The identified markers, particularly, OPA-07 and JH47, can serve as valuable tools for future studies aimed at exploring and exploiting the genetic diversity within the chrysanthemum population.

Standardization of in vitro micropropagation of Winter Jasmine (Jasminum nudiflorum) using nodal explants.

Present investigation was carried out to arrive at an effective micropropagation protocol for Winter Jasmine (Jasminum nudiflorum) using nodal segments from actively growing plants as explants. Explants were collected from current season shoots during April-May just after the initiation of new flush. Combined sterilization treatment of explants with 1.0% NaOCl2 for 10 min followed by 70% ethanol for 10 s recorded highest culture survival (63.88%) and optimum culture asepsis (63.88%) followed by the treatment containing 0.1% HgCl2 for 10 min followed by 70% ethanol for 10 s with culture survival (61.11%) and culture asepsis (69.44%). Highest culture establishment (80.55%) and minimum days to bud sprouting (7.62 days) was recorded with Benzyl adenine + Kinetin (3.0 + 1.0 mgL-1) but maximum length (4.33 cm) and leaf number (7.78) of established micro shoots was recorded with Benzyl adenine + Kinetin (1.0 + 0.5 mgL-1). Maximum proliferated shoots (2.41) and an optimum proliferation percentage (77.78 %) was recorded with Benzyl adenine + Kinetin (3.0 + 0.5 mgL-1). Minimum size of proliferated shoots (2.02 cm) was recorded with Benzyl adenine + Kinetin (3.0 + 1.0 mgL-1) followed by 2.25 cm recorded with Benzyl adenine + Kinetin (3.0 + 0.5 mgL-1). Highest rooting (63.93%), primary root number/microshoot (4.74) and longest primary roots (34.67 mm) were recorded with IBA (2.0 mgL-1). IBA yielded better results than NAA in terms of higher rooting percentage and root number. However, days to root initiation were found minimum (22.00) with 2.0 mgL-1 of NAA. Highest ex vitro survival of rooted microshoots (89.67%) was recorded with IBA (2.0 mgL-1).

Optimization of an improved, efficient and rapid in vitro micropropagation protocol for Petunia hybrida Vilm. Cv. "Bravo".

An efficient protocol for in-vitro propagation of an important ornamental crop, Petunia hybrida Vilm. Cv. "Bravo" was developed. The explants that were used to carry out the experiment were Leaf segments, nodal segments and shoot tips. Nodal segments recorded highest per cent asepsis followed by shoot tips and leaf segments. Asepsis was found to be highest when the explants were sterilized with Fungicide (Carbendazim) 0.02% for the duration of 30 min followed by 0.1% HgCl2 for duration of 10 min and then ethanol 70% for 10 s. Longer duration of the sterilant treatment showed more necrotic effects on the explants, thus mercuric chloride treatment when given for 5 min proved to be more effective in terms of survival of the explants. Maximum establishment per cent was recorded in Murashige and Skoog (MS) media fortified with BAP (1.5 mg L-1) and IBA (0.5 mg L-1) in shoot tips and nodal segments, i.e. 97.90 and 95.74% respectively. Callus was efficiently induced and developed when PGR amalgamation of BAP (0.1 mg L-1) and 2,4-D (1.5mg L-1) was used. Kinetin at the concentration of 2.0 mg L-1 along with IBA at 0.5mg L-1 recorded highest callus regeneration in both leaf and internodal segment derived callus. Maximum proliferation percent of shoots (97.90%), highest number of shoots (20.50 explant-1) and maximum length of shoot (2.70 cm) was recorded in PGR combination of IBA and BAP both at 0.5 mg L-1 concentration level. Rhizogenesis was recorded to be highest in the MS media containing IBA 1.00 mg L-1. Best hardening media which recorded maximum survival per cent 92.50% was noticed on the media formulation comprised of equal ratio of perlite and vermiculite mix, under poly house conditions.

First report of Fusarium solani associated with the bulb rot of Tulip (Tulipa spp) in India.

Tulip is an ornamental bulbous flowering crop belonging to the Genus Tulipa and family Liliaceae. It is the first ranking bulbous ornamental plant in the world (Nayeem and Qayoom 2015). They are often the first flowers to witness the bloom in the spring. Kashmir valley is located in northern Himalayas in northwestern region of Indian subcontinent. It is the most alluring and fascinating place all over India and the home of famous "Indhra Gandhi Memorial Tulip garden", the largest tulip garden in the entire Asia. However there are number of constraints in tulip cultivation among which bulb rot occupy a prominent place (Piwoni 2000). Bulb rot is posing problem to all the tulip growers throughout the world (De Hertogh et al. 1983). Rot symptoms were observed on tulip bulbs in field as well as in storage conditions (20-22◦C temperature with a relative humidity of 65%) in the summers of 2018 and 2019 in Shalimar fields of Kashmir. The main disease symptoms are yellow sunken spots on bulbs, purple-yellow coloration of leaves. Causal agent was isolated using tissue bit technique (Pathak 1972) on potato dextrose agar plates which where incubated at 24±2◦C . Single spore technique was used to obtain the pure isolate (Johnston and Booth 1983). The isolate covered the full plate (90mm) in ten days. The colony was dull whitish in color, flat and smooth with concentric ring formation in the culture plate with inner ring having a creamy exudation. The mycelium was septate, branched and hyaline in color and measured 3.50-5.20 µm in width with an average of 4.4 µm. Micro-conidia were hyaline, cylindrical to oval, 0-1 septa and measured 7.50-11.00×2.80-3.75 µm in size. Macro conidia were hyaline with 3-4 septa, fusiform, moderately curved which measured 21.15- 32.00×3.80-4.75 µm in size with an average of 28.50±0.21× 4.30±0.2 µm. On the basis of these morphological and cultural characteristics of the fungus, it was identified as Fusarium solani (Mar.) Sacc.,. To confirm the identity the PCR amplification was carried out for two genes Internal Transcribed Spacer (ITS 1, ITS 4)and Translation Elongation factor1-alpha gene (tef1- alpha) (O'Donnell et al. 1998; White et al. 1990). BLAST analysis of the sequence obtained for both the genes showed 99% homology with F. solani sequences in GenBank and Fusarium -ID databases. The sequences were deposited in the GenBank (Accession No MN611433, MW995477). Pathogenicity test was conducted on variety orange emperor both in laboratory and polyhouse. Bulbs were divided into three sets, (three bulbs per set) one set was given injury and dipped in conidial suspension (106 conidia/ml) for 30 min, another set was kept uninjured and dipped in spore suspension of same concentration, the third set was served as control and dipped in sterilized distilled water. All the respective sets were incubated in a moist chamber maintained at a temperature of 22 ◦C to observe symptoms. The injured ones showed symptoms after 7-8 days of inoculation, whereas the uninjured bulbs showed symptoms after 11-12 days. No symptoms were observed in controlled set. A pot experiment was also conducted to carry the pathogenicity tests. Bulbs were injured with the help of sterile needle and were dipped in conidial suspension (106 conidia/ml) for 30 min (Pastrana et al. 2014). The bulbs kept for control were dipped in sterilized distilled water. Bulbs were then planted in pots maintained at 18◦C. The above ground parts of the inoculated bulbs showed symptoms like stunted growth which gradually turned yellow and did not produced flowers. The bulbs after harvesting were rotten .No symptoms were observed in controlled plants. To fulfill the Koch's postulates the fungal pathogen was re-isolated which was identified as F. solani. The pathogen is reported to cause disease in other crops (Gupta et al. 2012) but to our knowledge and on the basis of literature, this is the first report of F. solani causing bulb rot of tulip in India.