Comparative Analysis Of Anatomy And Cell-cycle Related Genes Of Southern Highbush Vaccinium Corymbosum

Cell-cycle system controls the flower and fruit growth,size and weight precisely,accompanied by cumulative cell proliferation and cell expansion.Blueberry is an evergreen woody plant of the genus Ericaceae(Vaccinium spp.),which is usually divided into 21 groups and 450 species.Vaccinium corymbosum shows significant variation in berry size,but the molecular mechanism regulating the size of the fruit is still unknown.Therefore,in this study,single fruit weight,cell layers,cell number and cell area of Southern highbush blueberry cultivars ’O’Neal’ and ’Bluerain’ during V.corymbosum flower and fruit during ontogeny were analyzed.The sequences of VcCyclins and VcCDKs were isolated,and its expression tendency was also detected.The specific results are as follows:(1)The fruit growth of blueberry exhibited a double-sigmoid pattern,and flower buds had a lower contribution to mature fruit weight and while ovary wall(or pericarp)was the vital component for final fruit size and approximately 70% cell layers of pericarp were produced before anthesis.(2)The cell number exhibited obvious changes in the different tissues of blueberry fruit:(1)The number of epidermis cells increased in the whole fruit development,but showed two rapid growth periods.(2)Endocarp cells were mainly divided in the stages I-S1 until the late development stage,the endocarp cell number decreased slightly.(3)The cell numbers of outer mesocarp was only increase at stages III-S1,and the second rapid growth period(S3-S4).(4)Cell numbers of mesocarp and inner mesocarp only increased slightly in the early stages of flower bud enlargement(I-II or I-III),then remained unchanged.(5)As placental tissue,the cell numbers continued increasing at a low rate until stage IV,and decrease dramatically from stage S2 or S3.(3)Cell area changes of different parts of blueberry fruit:(1)The cell area of epidermis and hypodermis maintained a low growth rate in the whole flower bud enlargement and fruit development,(2)While endocarp cells mainly expanded during the fruit development(S1-S6).(3)The cell area of outer mesocarp,mesocarp and inner mesocarp continued to expand at a low rate throughout the flower bud enlargement,but increased rapidly during the fruit development.(4)The incrase of placental tissue cell area was relatively slow in the stages I-S3,and increased significantly in the stages S3-S6.(4)Compared cell numbers and cell area between ’O’Neal’s and ’Bluerain’s,the cytological difference of two cultivars was as follows:(1)The numbers of epidermis and hypodermis cells of ’Bluerain’ were lower than ’O’Neal’ from stage I to fruit ripening,and the single cell area of ’O’Neal’ was also larger than ’Bluerain’.(2)The number of endocarp cells in ’O’Neal’ was larger than ’Bluerain’ in the whole fruit development process,but there was no significant difference between the two cultivars.(3)The number of outer mesocarp cells in ’O’Neal’ from stage IV was more than ’Bluerain’,and showed three rapid growth stages: stages III-IV,stages S0-S1 and stages S2-S4.The single cell area of outer mesocarp in ’Bluerain’ was larger than ’O’Neal’ in I-IV and S2-S6 stages,which was also the fastest expansion period for outer mesocarp cells in ’Bluerain’.(4)The cellnumber of middle mesocarp and Inner mesocarpin ’O’Neal’ increased rapidly from the early stage of flower bud enlargement to fruit ripening,whilethecellnumber in ’Bluerain’ maintained constantly during the whole development process,except increased slightly at the fruit ripening stage.(5)The number of ’O’Neal’s placenta cells began to decrease from stage S2 and was slightly higher than ’Bluerain’ at fruit ripening stage,and the cell area in ’O’Neal’ was larger than ’Bluerain’ during the whole flower and fruit development.From stage S0,’O’Neal’s growth rate began to increase significantly,while ’Bluerain’s growth rate of placenta area increased significantly from stage S3.The results showed that the number and area of cells in the outer mesocarp and placenta were the key factors affecting fruit size,and the critical period that affects fruit size between ’O’Neal’ and ’Bluerain’ might be earlier from the first stage of flower bud enlargement.(5)12 cell-cycle related genes were isolated and detected from ’O’Neal’ flower buds by homologous cloning.The sequences of VcCyclins and VcCDKs were relatively conserved.The expression patterns of VcCycA2 L and VcCycB2 L in flower bud stage II-IV and III-V were higher in two cultivars,respectively,the expression trend of VcCDKB2;2 was consistent with VcCycA2 L and VcCycB2L;VcCycD3;1 and VcCycT1;4L expressed abundantly in the stage S0-S4,but the expression pattern of VcCDKE1 tended to be consistent;the expression patterns of VcCycP3;1L and VcCDKC1 L increased and decreased regularly during the development of flower buds and fruits;VcCycH1;1,VcCDKC1 L and VcCDKG2 L may not directly participate in fruit growth and development.In addition,there was no significant difference in the expression of these genes in the process of flower bud enlargement and fruit development,suggesting that the 12 cell cycle-related genes isolated were not the key regulatory genes affecting fruit size of ’O’Neal’ and ’Bluerain’.