Identification Of Different Wild Rice And Construction Of Molecular Marker Map Of O.Eichingeri

The genus Oryza consists of more than 20 species,including about 20 wild Oryza species and 2 cultivated sub-species.The wild Oryza species have been classified into 11 distinct groups(A,B,BC,C,CD,D,E,F,G,HJ and HK).To study the possibility of application of oligonuclitide probes in wile rice and evolutionary relationships of different wild rice,we conducted fluorescence in situ hybridization(FISH)using 45 S rDNA and a library containing 25,000 oligonucleotides(oligos)probes in different wild rice.The pool of the oligos(oligo-chr9)had developed from the sequence of chromosome 9 of a cultivated rice,Nipponbare.The main results are as follows:1.There are 24 chromosomes in each somatic cell of Oryza punctata(YS7020,BB).There are 2 oligo-chr9 and 4 45s rDNA probe signals visualized by FISH.A pair of 45s rDNA signals located on chromosome 9 are strong,another pair signals are weaker.Oryza minuta(YS7021,BBCC)contains 48 chromosomes.Four oligo-chr9 and 4 45s rDNA probe signals were observed.However,no 45s rDNA probe signal was located on the end of chromosome 9.Oryza alta(YS7035,CCDD)has 48 chromosomes in each somatic cell.There are 4 oligo-chr9 and 8 45s rDNA probe signals in each cell visualized by FISH.Only 3 signals of 45s rDNA are located on the ends of chromosome 9,one distributed on the end of other chromosome.The other four 45S rDNA signals are located at pericentromeric region of the other chromosomes.One line of O.latifolia(YS7036,CCDD)has 48 chromosomes in each cell.Four oligo-chr9 probe signals were observed in each cell.Two of the oligo-chr9 probe signals were weaker and the another two were stronger.There were 14 signals of 45S rDNA probes.Eight signals of them located on the ends of chromosomes(four of them are chromosome 9).The remain 6 signals of 45S rDNA distributed at pericentromeric region of the other chromosomes.Eight 45S rDNA signals were observed in the other line of O.latifolia(YS7037,CCDD).Four of them located on the ends of chromosome 9.Two of 45S rDNA signals located on the ends of other chromosomes and 2 of them located on pericentromeric regions of the another chromosomes.Oryza brachyantha(YS7040,FF)has 24 chromosomes.Two signals of 45S rDNA distributed at the end of chromosome 9.Oryza longiglumis(YS7039,HHJJ)has 48 chromosomes.Four signals of 45S rDNA were distributed at the ends of chromosome 9.Two signals of them were located at the ends of the other chromosomes.And the other two signals of 45S rDNA distributed at the pericentromeric region of the another chromosomes.Oryza ridleyi(YS7042,HHJJ)has 48 chromosomes.Four signals of 45S rDNA located on the ends of chromosome 9.2.There are different species in the wild rice with CC genomes,including O.eichingeri,Oryza rhizomatis and Oryza officinalis.We found that 2 oligo-chr9 and 2 45S rDNA signals in the line YS7027 of O.rhizomatis.One of the 45S rDNA signals was distributed at the end of chromosome 9,and the another one was distributed at the end of the other chromosome.Different lines of O.eichingeri and O.officinalis have different 45S rDNA and oligo-chr9 probe signals.The karyotypes of different lines of O.eichingeri and O.officinalis showed that differentiationts have occurred.Such as in O.eichingeri(YS7024 YS7025 YS7026),45S rDNA probe signals appeared as 5,3 and 2 in YS7024,YS7025 and YS7026,respectively,and only one 45s rDNA located at the end of chromosome 9.In O.officinalis(YS7029、YS7030、YS7032),differences exist in both distributions of 45S rDNA and oligo-chr9.Such as oligo-chr9 signals distributed on 2 chromosomes in YS7029 and YS7032.But it distributed on 4 chromosomes in YS7030.Two pairs of 45S rDNA probe signals located at the ends of 4 chromosomes in YS7029 and YS7030.However,additional pairs of 45S rDNA probe signals distributed at pericentromeric region of 2 chromosomes.3.According to the sequences of the centromere of O.officinalis,5 pair primers were selected.One pair primer gave specific target bans in the three types wild rice with CC genomes,0.eichingeri,0.rhizomatis and 0.officinalis.After cloned to the vector,the target sequence,CentO-C was labeled as probe by nick translation.FISH result showed that there are 20 signals in each cell in 0.eichingeri,O.rhizomatis and 0.officinalis.Two pairs of chromosomes had no signal of CentO-C.It is a useful tool to detect chromosomes derived from the wild rice with CC genome.4.The 352 pairs of SSR polymorphism molecular markers located on 12 chromosomes between 0.eichingeri and cultivated rice 02428 were selected and a molecular marker map was constructed.It provides a basis for the utilization of of O.eichingeri.