Distinguishing Responsive Microspore Developmental Stages and Inducing Stressors for Androgenesis in Sweet Sorghum

Aron Felts, Tennessee State University


Sweet sorghum is a promising crop for food, feed, and biofuel production. The objective of this study was to assess the potential of individually isolated immature pollen cells, also called microspores, of sorghum to be induced to produce embryogenic calli allowing for double haploid production. Two varieties of Sorghum bicolor L. (Dale and Topper 76-6) were grown in a greenhouse to record plant height, flag-stem length as well as spikelet maturation characteristics at the booting stage for capturing microspores. Greenhouse data was correlated with microspore developmental stages to harvest anthers containing immature free microspores after measuring spikelet and anther sizes. This allowed accurate harvesting of panicles containing six stages of microspore development, i.e., tetrad, early unicellular, late unicellular, early bi-nucleate, mid bi-nucleate, and mature pollen. Under development of exine in isolated microspores was confirmed through various cell staining protocols as well as REPLI-g based single cell DNA amplifications. Anthers were dissected for mass isolation of microspores through centrifugation which was then washed with sugar gradient to eliminate excess debris and dead cells. Early unicellular to mid bi-nucleate microspores were cultured onrich media causing maturation after one week and survival up to one month or longer at 4 °C. Topper 76-6 panicles containing immature microspores were subjected to 21-day cold pre-stress treatment for the induction of androgenesis. Microspores were plated on various concentrations of growth regulators to test for androgenesis responsiveness. MS salts with 5 mg/L 2,4-D and 2mg/L kinetin lead to production of compact calli, while media with no hormones gave friable calli.

Subject Area

Plant sciences|Cellular biology|Developmental biology|Agriculture

Recommended Citation

Aron Felts, "Distinguishing Responsive Microspore Developmental Stages and Inducing Stressors for Androgenesis in Sweet Sorghum" (2020). ETD Collection for Tennessee State University. Paper AAI28027885.