Micropropagation usually involves cytokinin in single-harvest batches. We report two in vitro studies with multiple harvest: (1) fed batch process with modified physical states and (2) LED light treatments. In (1), genotypes of Cannabis sativa were observed in stationary agar (A), stationary Oasis® infused with liquid (OIL) and agitated Oasis® infused with liquid (AOIL).Fifteen explants were planted in vessels with 120 mL DKW medium harvested on 3-week cycles, with 0 or 15 mL additional media. Harvested shoots, length, and dry mass from repeated cycles were recorded. Genotypes T1 and Peach failed on multiple harvest cycles and were eliminated, although single cycle had higher quality in OIL. BaOx and Cherry1 on OIL/AOIL with additions were better quality than A in five cycles. Shoots harvested increased from 15 to 30 in cycles 1-3 in OIL/AOIL, but in A were approximately 20, while length was longest in OIL/AOIL. By cycle 3, all measured responses were decreasing until cycle 5 where a minimum of 7 shoots per vessel or more were only in OIL, but shoots were too short to plant in greenhouse. In (2), blue and supplemental far-red were observed with in vitro shoots of BaoX and Cherry1. OIL treatments were placed in LED polychromatic and dichromatic light (white, high red:blue, medium red:blue, white w/5% far-red, high red:blue 5% far-red, medium red:blue w/5% far-red, low red:blue w/ 5% far-red) at similar intensities (190-240 µmols·m-2·s-116 h-photoperiod). Media additions were made with responses recorded bi-weekly. Five randomly selected microcuttings per vessel rooted ex vitro on mist bench for 16 days. Over multi-cycles, plants treated with 5% far-red increased number and length, while plants under higher blue light increased dry mass. Shoot number increased to 28 in cycles 1-3 with far-red, and 18 without before decreasing to initial 15 during cycle 5. The accumulated shoots per vessel over 5-cycles (10-weeks) was 108 with far-red, and 84 without. Shoot length in far-red-treated plants increased from 19 - 25 mm in cycle 3 before decreasing to 10 mm in cycle 5. Plants without far-red had 10 – 15 mm length the entire experiment. Dry mass was highest during cycle 1 with blue light before decreasing 50% in cycle 3, where it remained until cycle 5. Sixty-eight percent of shoots rooted regardless of prior in vitro treatment. OIL with media addition allowed shoots to be harvested five cycles, while signaling response of far-red light allowed increased productivity and length.
