Study on aseptic sowing technique of Orchid Seeds

1, capsule treatment 1, selection, pretreatment to select the capsule after pollination for a certain number of days, generally take the capsule with yellowish skin and dull pericarp. For the recovered capsule, first rinse the surface with obvious dirt such as sediment and dust under tap water, then wipe the surface and groove seam with cotton dipped in diluted detergent solution, then rinse under tap water for 15-30 minutes, and cut off the front segment with an anatomical knife (be careful not to cut too much to prevent capsule cracking). 2. After the capsule is dried after surface disinfection and rinsing, it depends on the situation on the super-clean worktable. a. When the capsule is large and the pericarp is thick and hard, soak in 75% alcohol for a few seconds to a few minutes, then remove, burn and sterilize, disinfect the surface and burn dry. b. When the capsule is small, soak it in 75% alcohol for 5-10 s, then disinfect 15-20min with 0.1% mercury from Tween, then rinse the capsule with sterile water 4-5 times, 1-2min each time, and remove it in sterilized Kraft paper to dry. Second, sow the sterilized capsule on the ultra-clean worktable on a sterile cushion plate, tweeze the capsule with sterilized tweezers, cut off both ends of the sterilized capsule with an anatomical knife (you can see the powdery seeds), then plane the capsule from the hook seam, scrape off the seeds in the capsule, and spread evenly on the surface of the medium. 3. Induction culture 1. Huabao No. 1 and MS medium were often used in the culture medium, and activated carbon, sucrose and Agar were added. According to the different proportion of 6 murine BA and NAA, pH was adjusted to 5.4-6.0. 2. The culture temperature is about 26 ℃, and there may be scattered light in the early dark culture. After the protocorm grows obviously, the light intensity is about 2000 Lx, 12h/d. After a certain day of sowing, the seeds began to germinate after being cultured on the induction medium for a certain number of days, but the embryo did not elongate, but the embryo gradually expanded. When one end of the seed coat was broken, the swollen embryo was yellow, similar to callus, and continued culture. the surface of the embryo is covered with pseudo-protocorm bodies of many small cones. With the extension of culture time, the small conical sphere gradually expanded to form a green spherical protocorm, which can be used for proliferation, subculture and differentiation. 3. After subculture, after sowing for a certain period of time (about 50-60 days), the original bottle space of induced culture is almost full after the seed germinates large enough protocorm, in order to increase the growth space, the bottle can be divided. When dividing bottles, the seedlings should be separated as far as possible and evenly distributed in the medium. It is common to separate bottles every (60-80d). 4. Rooting culture of strong seedlings after several times of subculture, some protocorms gradually grow into seedlings with 1-2 leaflets, and some seedlings have 1-2 thick and short green roots. at this time, these seedlings can not be directly refined and transplanted, and then the shaped seedlings can be cultured. The varieties and plant types of Ophiopsis were added to the rooting medium of strong seedlings such as banana paste and coconut juice, and continued to be cultured until they grew into seedlings with 3-5 leaves, 3-4 roots and high 5cm, and then transplanted and domesticated. The culture temperature is about 26 degrees, the early light intensity is about 2000 Lx, and the light time is 12h/d. The scattered light of sunlight can be used in the later stage, and the light intensity is (800-1000) Lx.