| The genus “Carya” belongs to Juglandaceae family.This genus contains important economic tree species with significant economic value,among which Carya cathaynesis Sarg.and Carya illinoinensis are two economically important species that playing important roles in the nut industry of the Zhejiang Province.In recent years,the demand for elite varieties of hickory are growing due to the rapid development of hickory industry.However,the supply for them is far from meeting the requirements because the genetic breeding of hickory develops very slowly.The genus Carya contains about 16 species with disjunctive distributions,among which 11 are distributed in Northern America and 5 in East Asia.Different Carya species displayed significant variations in terms of tree height,resistance,yield and quality,thus it is necessary to breed among different species for the desired characteristics.To date,the genome size,genotype and karyotype of most Carya species,as well as the phylogenetic relationships between them are still unknown,which have become major limitation for developing the crossbreeds among different Carya species and the molecular biology research of Carya species.In this study,ten Carya species,including C.cathaynesis Sarg.,C.tonkinensis Lecte,C.hunanensis,C.dabieshanensis,C.illinoinensis,C.ovata,C.myristiciformis,C.aquatica,C.texana and C.floridana,were selected as study material and investigated for their genome size,karyotype and physical distribution of 45 S rDNA through flow cytometry,karyotype analysis and fluorescence in situ hybridization(FISH).K-mer analysis was performed to evaluate the percentage of repeated sequences in the genomes of C.cathaynesis Sarg.,and C.illinoinensis based on their sequencing data.The results are as follows:(1)According to the differences in chromosome number,10 Carya species were divided into 3 types: The first type contained species with the chromosome number of 2n=32,which included C.cathaynesis Sarg.,C.tonkinensis Lecte,C.hunanensis,C.dabieshanensis,C.myristiciformis,C.aquatica,and the karyotype formulas of these species were 2n=32=20m+12sm,2n=32=16m+16sm,2n=32=22m+10sm,2n=32=22m+10sm,2n=32=26m+6sm and 2n=32=22m+10sm,respectively.The second type were the species with the chromosome number of 2n=34,mainly including C.illinoinensis and C.ovata with the karyotype formulas of 2n=34=22m+12sm and 2n=34=24m+10sm,respectively.The third type were the species with the chromosome number of 2n=64,including C.floridana and C.texana,and their karyotype formulas were 2n=64=52m+12sm and 2n=64=42m+22sm,respectively.The chromosome types of these ten Carya species were median region chromosomes and submedian region chromosomes.The karyotype type of these ten Carya species belonged to 2B.These results indicated that the karyotype of these Carya species were relatively symmetrical.(2)The chromosome behavior of most pollen mother cells in C.cathaynesis Sarg.,is normal and paired as 16 bivalents at metaphase?.However,there is a certain proportion of abnormal pollen mother cells in meiosis,such as appearing lagging chromosome in metaphase I,which accounts for 8.86%,showing asymmetry division in metaphase II,which accounts for 15.79%.(3)Flow cytometric analyses showed that the genome sizes of C.floridana and C.texana were(1193.947±3.093)Mb and(1017.751±6.486)Mb,respectively,which were significantly larger than those of the other 8 diploid Carya species.Among the 8 diploid Carya species,C.cathaynesis Sarg.had the largest genome size of(727.330±5.373)Mb,followed by C.illinoinensis,with the genome size of(690.939±6.969)Mb.C.ovata had the smallest genome size of(588.957±9.527)Mb.Among the 6 diploid Carya species with the chromosome number of 2n=32,C.hunanensis and C.dabieshanensis had more similar genome sizes of(671.359±4.423)Mb and(687.538±3.507)Mb,respectively;C.myristiciformis and C.aquatica had similar genome sizes of(607.323±3.357)Mb and(606.533±9.357)Mb,respectively.C.cathaynesis Sarg.and C.illinoinensis had different repeat sequence percentages of 57.3% and 48.2%,respectively.(4)As for the difference in the number of nucelotides per chromosome,among the ten Carya species,C.hunanensis and C.dabieshanensis had the smallest difference,for 10 pairs of chromosomes between which the difference was less than 2M.For 9 pairs of chromosomes between C.myristiciformis and C.aquatica,the difference was less than 2M.However,there were more difference between C.cathaynesis Sarg.and C.myristiciformis,C.cathaynesis Sarg.and C.aquatica,C.illinoinensis and C.Ovata,as well as C.floridana and C.texana.(5)According to the characters of 45 S rDNA distribution,10 Carya species could be classified into two categories: The first category was the species with only one 45 S rDNA distribution type,including C.cathaynesis Sarg.,C.hunanensis,C.dabieshanensis,C.myristiciformis,C.aquatica,C.texana and C.floridana.The other category was the species with a variety of 45 S rDNA distribution type,including C.tonkinensis Lecte,C.illinoinensis and C.ovata.Among which,C.tonkinensis Lecte had three different 45 S rDNA distribution types,C.ovata had two types and C.illinoinensis had five types.(6)Compared to other species,45 S rDNA distribution of Carya species was complex.Based on the loci number and location,45 S rDNA distribution of ten Carya species could be divided into three types.For the first type,the loci number of 45 S rDNA was even and 45 S rDNA was distributed in pairs on homologous chromosomes.For example,C.dabieshanensis had one pair of 45 S rDNA loci on its homologous chromosomes;C.myristiciformis,C.aquatica,C.texana and C.floridana had two pairs of 45 S rDNA loci.For the second type,the number of 45 S rDNA loci was even,but there were differences in signal intensity and distribution locations.For example,for the two 45 S rDNA of C.hunanensis,one was located at the end of chromosome,the other was located in the center of chromosome arm;some 45 S rDNA in certain cells or individuals of C.tonkinensis Lecte and C.illinoinensis were located in different chromatids.For the third type,the number of 45 S rDNA loci was odd.For example,the number of 45 S rDNA loci was three in C.cathaynesis Sarg.and some cells or individual of C.tonkinensis Lecte,five in some cells or individual of C.illinoinensis and C.ovata. |
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