Black Rice (indica type) Originated from the Wild Rice of India During Domestication: Genetic Evidence from Interspecific Hybridization Utilizing Non-Black Parents (O. sativa x O. rufipogon)

I have developed black rice RIL lines from the interspecific hybridization between non-black parental lines (Oryza sativa and O. rufipogon) in 2016. Black rice RIL lines BWF (Badshabhog x O. rufipogon) and another second black rice breeding line CWF (Chenga x O. rufipogon) were analyzed for the black pericarp trait which was inherited in the progeny lines (F2, F3, F4, F5, F6, F7, F8, F9) without any morphological constrains signifying that black pericarp related genes were fixed. Parental lines were non-black such as rice cultivar Badshabhog is white grained, Chenga is brown grained and wild rice O. rufipogon is red grained. Three genes Kala1, Kala3 and Kala4 are responsible for black pigmentation in rice (Maeda et al. 2014; Oikawa et al. 2015). So, in this present investigation, possible reason for black pericarp formation in the breeding lines may be that the LINE1 insertional rearrangement mutation in Kala4 gene (Oikawa et al. 2015) or other unknown new and novel genetic changes have been occurred leading to ectopic expression of anthocyanin biosynthesis genes and ultimately gave birth of black rice in the breeding lines (acquired neo-functionalization). Mendelian genetics and phenotypic characteristics still at the F9 generation (2024) confirming our model- Black Rice (indica type) Originated from the Wild Rice of India During Domestication: Genetic Evidence from Interspecific Hybridization Using Non-Black Parents (O. sativa x O. rufipogon). Genome-wide variants (SNP/InDel/SV) analysis was conducted using long read Nanopore sequencing technology in six rice lines Chakhao_ML (control black line), wild rice O. rufipogon (Or_Rai), local cultivars Badshabhog, Chenga, and two black breeding lines BWF107 and CWF163 to understand the genome-wide variations specifically to novel traits black pericarp. Unique variants (SNP/InDel/SV) were identified for the genes associated to black pericarp formation Kala1 (Os01g0633500), Kala3 (Os03g0410000), Kala4 (Os04g0557200, Os04g0557500, Os04g0557800) and many other genes. These unique variants may be responsible for the chromosomal rearrangement to acquire gain-of-function mutation (neo-functionalization) leading to black pericarp formation in the breeding lines (BWF, CWF). Interspecific hybridisation with non-black parents (O. sativa x O. rufipogon) provided genetic evidence that domesticated black rice (indica type) originated from wild rice of India. Hence, underutilized wild rice (O. rufipogon) contributed immensely to enhancing the genetic base of the breeding lines to uncover the hidden genetic construct related to black pericarp pigmentation and our novel black rice breeding lines (RIL) may provide some insight into the evolutionary history of black rice origin.