Cloning And Expression Profile Analysis Of Key Genes CsHd3a And CsCoL Gene Family In Cannabis Flowering

Photoperiodism has a great interaction with plant flowering.Many complicated networks are regulated by light signaling.CO genes are the key factor in the regulation via biological clock processes.The upstream CO genes sense the signal and transmit to the downstream FT genes,activating flowering organ differentiation and growth.Based on the theoretical basis of previous studies on flowering-related genes in plants such as Arabidopsis,rice,cotton,et al.This study investigated the flowering period of179 cannabis germplasm resources in the field under short-day(SD)condition,and selected representative early/late flowering varieties as research materials,using bioinformatics and molecular biology and other technical methods to study the key genes of cannabis flowering CsHd3 a and CsCOL family genes.Using bioinformatics methods,gene sequences were obtained from cannabis genome data,fragments were cloned by PCR technology,and real-time fluorescence quantitative PCR(qRT-PCR)technology was used to explore the temporal and spatial expression of genes.The results as follows:1.The flowering period and budding period were investigated on 179 cannabis germplasm resources.Most of the varieties in northern China showed early flowering after planting in southern China,that is,buds appeared 19 to 35 days after sowing,and they bloomed 23 to 38 days after sowing.Southern varieties such as "Yunma" and "Bama" series are mainly focused on budding after 50 days and blooming after 63 days.The variety "668" from Jilin,"Yousha 300" from Heilongjiang and "Lvliangma" from Shanxi are all northern varieties,but its flowering time is 56 days after sowing,which is the same as that of some southern varieties,indicating that these varieties are less affected by the photoperiod after they move from the north to the south,and they can be used as candidates for wide adaptability for breeding.2.The CsHd3 a gene was cloned in the cannabis variety "Q1",with a total length of 543 bp,encoding 180 amino acids containing the PEBP family protein domain.Tissue-specific analysis showed that the gene was expressed first in the leaves and showed rhythmic changes under SD treatment,culminating at 8 a.m.Further analysis showed that the expression of CsHd3 a gene in the early flower variety "Q1" was significantly higher than that of the late-maturing variety "Y7",especially during the day.In addition,there are several amino acid differences in the CsHd3 a amino acid sequence in "Q1" and "Y7".To sum up,CsHd3 a encodes a member of the PEBP protein family and may be involved in cannabis light cycle flower regulation.3.A total of 13 CsCOL genes were identified in the cannabis genome.Systematic developmental analysis with amoeba and rice showed that the CsCOL protein was divided into three subgroups,each containing a conservative intrinsic/exon structure and base sequence.Chromosomal location analysis showed that 13 CsCOL genes were unevenly distributed on 7 chromosomes,of which CSCOL members were the largest in chromosome 10.Colinear analysis showed that homologous genes CsCOL4 and CsCOL11 were found in rice and Raymond cotton.Of the 13 CsCOL genes,CsCOL6 and CsCOL12 are a pair of concatenation repeat genes,and CsCOL8 and CsCOL11 may be duplicated in segments.In addition,tissue-specific expression indicates that 10 CSCOL genes are expressed first in the leaves,one in the stem and two in the female.Most CSCOLs exhibit day and night fluctuations under different light cycles.In addition,CsCOL3 and CsCOL7 have several amino acid differences between early-flowering and late-flowering varieties.