| Secale cereale L.cv.Jingzhouheimai(JZHM)is an important genetic resource for powdery mildew resistance improvement of wheat.Previously,after RNA-seq,total 70,705 non-redundant sequences were generated and 46,810 contigs were obtained via de novo assembly from Bgt(Blumeria graminis f.sp.tritici)induced seedlings of JZHM.From them,3,197 up-regulated genes were identified.In this study,distribution of SSR was analyzed,molecular markers specific for rye chromosome were developed and chromosomally mapped,specific markers were also used in alien chromosome lines selection,the main results were as follows:Using MISA program,SSR distribution in 70,705 non-redundant sequences was analyzed at the website http://pgrc.ipk-gatersleben.de/misa/,total 16,697 SSRs were identified in 11;904 contigs(16.84%),the average distribution rate of SSR was one per 2.37 kb.8,660(72.75%)contigs contained only one SSRs,while 3,244(27.25%)contained more than one SSRs,and 2,240 SSRs occurred in compound forms.Among these SSRs,trinucleotide motifs(9,593,57.45%)were the most,in which motifs CCG/CGG(3,289,19.70%)were the most.According to SSRs length,12 bp SSRs were the most(9,810,58.75%),followed by 15 bp SSRs(3,090,18.50%),the number of SSRs longer than 25 bp was only 31(0.20%).According to 3,197 up-regulated genes,294 new primers pairs were designed,combined with 343 and 237 pairs of primers designed by Liu(2015)and Ding(2015)respectively,total 874 primer pairs were used to identify specific markers which included 176 pairs of SSR primers(class I),148 pairs of PLUG primers(class ?),237 pairs of primers designed on no similar sequences against wheat D genome(class ?)and 313 pairs of primers designed on no similar sequences against NCBI non-redundant nucleotide database(class IV).After amplified in HXH(T.aestivum landrace Huixianhong),JZHM and JH 1(HXH-JZHM amphiploid Jinghui 1),besides 88 without amplification,786 produced stable amplicons,of them 307 primer pairs produced 403 rye specific loci,which included 87 primers of class I,42 primers of class II,80 primers of class ? and 98 primers of class ? with the polymorphic rate at 39.06%.Using a set of HXH-JZHM alien chromosome lines involving 7 chromosomes,168 markers were mapped on different JZHM chromosomes,141 markers were mapped on one rye chromosome,10,22,17,39,19,25,and 9 markers were respectively located to chromosomes 1R to 7R,and the other 27 markers were duplicated on more than one chromosome.These markers were further used to identify rye chromosome aberrations.Among them,nine markers(including 2 markers previously reported)specific for chromosome 5R were selected to analyze DtA5RS and a F4 population derived from CS phlbphlb/MA5R,6 plants with absence of 4-8 markers were identified,and one plant was characterized as a monnosomic addition MtA5RL after FISH analysis.Based on the amplification on these lines,the nine markers were allocated which included three on 5RS and six on 5RL.Additionally,combined molecular and cytological marker analysis,10 wheat-rye introgression lines involving chromosomes 1R and 7R were identified. |
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