Somatic hybridization for citus scion and rootstock improvement against greening disease and for seedlessness

Abstract

Citrus greening or Huanglongbing (HLB) is one of the most destructive diseases of citrus worldwide. It is threatening survival of the citrus industry in Pakistan. The causal agent of HLB is ‘Candidatus Liberibacter asiaticus’. The search for alternative citrus greening tolerant/resistant scion and rootstocks that also withstand the other constraints is needed for a sustainable citrus industry. The production of somatic hybrids allows accumulating all the dominant traits of both parents, irrespective of their heterozygosity level, and has proven to be an important tool for rootstock improvement. A new embryogenic callus line and suspension culture of the seedy and polyembryonic ‘W. Murcott’ mandarin hybrid has been developed. Citrus latipes (papeda) a monoembryonic citrus has been purported to exhibit tolerance to HLB. Somatic hybridization of ‘W. Murcott’ mandarin hybrid with Citrus latipes was conducted by protoplast fusion of embryogenic suspension culture-derived protoplasts isolated from ‘W. Murcott’ mandarin with leaf protoplasts of C. latipes. Analysis of ploidy level by flow cytometry confirmed the regeneration of thirty five new interesting tetraploid somatic hybrids from this combination. Furthermore, the morphology of the new putative somatic hybrids was intermediate between the parents, exhibiting the larger winged petiole and elongated leaf characteristics of C. latipes. Molecular markers analysis of the new somatic hybrids to determine the allotetraploid and autotetraploid hybrids showed that at least few hybrids are allotetraploid. Another fusion for scion improvement was performed between W murcott suspension protoplast and Snack mesophyll protoplast. Snack is a tangor hybrid and is showing greening tolerance up till now. Fourteen plants were regenerated from W murcott and Snack combination and all were confirmed as autotetraploids. W murcott is best regenerative parent and performs well under tissue culture conditions. It has drought tolerance. Its hybrids can perform well under arid conditions of Pakistan. Its breeding with Kinnow can produce triploid crop of an intermediate season. The new somatic hybrids will be propagated to evaluate their horticultural performance and ability to mitigate or prevent HLB in grafted commercial scions. The somatic hybrid rootstocks should also have potential to control tree size due to polyploidy, and should be amenable to standard seed propagation via nucellar seed production. W. Murcott was used as a parent in this project because it is a mandarin hybrid that is seedy and polyembryonic, a trait needed for traditional seed propagation of rootstocks. These hybrids will add new germplasm to the citrus scion and rootstock breeding program, potentially addressing the HLB problem.