| Walnut(Juglans regia L.),also known as walnut,?Qiangtao?,is Juglans Branch(Juglandaceae),Walnut(Juglans),is one of the four famous nuts.In recent years,the transcriptome sequencing technology in biological sequences apply in almost research field and widely recognized in biology genetic study.As one of the typical nuts,few studies in analysis of the transcriptome sequence,especially the molecular mechanism on formation of walnut endocarp.Therefore,in order to strengthen the research of walnut transcriptome information,understanding the molecular mechanism of walnut endocarp formation,grasp the main genetic information affecting the walnut endocarp,the walnut varieties of ?Zhi Pi?,?Wen 138? were used in the study,the endocarp were separated,then RNA were extracted,RNA-Seq and analysised the data,the main results are as follows:1.Use Illumina Hi Seq? 2000 sequencing instrument for sequencing,we acquire raw reads 342 785 132,after filtering,acquire clean reads 318 196 894 and the length of 28 637 720 460 nt.In order to obtain information under the condition of accuracy is 99%,98.6% of sequence information conform to the requirements of the analysis,the data obtained amount of information.After De novo assembly,acquire total unigenes 77 570,the total length of 97 085 606 nt,the average length of 1 252 nt,N50 is 1 978 nt,which N50 significantly greater than average length,it is better to illustrate the assembly.2.Unigene of splicing after assembly for functional annotation,compareing the sequence information with known database NR,NT,Swiss-Prot,KEGG,COG and GO,there were 56 139,53 032,37 712,34 325,22 517,44 620 unigenes sequence showed high homology with the above database,comparing to a total of 58 382 unigenes,accounting for 75.26% of the total unigenes,alignment effect is good,greatly enriched the walnut endocarp transcriptome of information.A number of SSR loci were finded in 18 885 unigenes,which included in 77 570 unigenes by using MISA software,accounting for 24.35% of the total unigenes,and 4 143 unigenes include more than one SSR loci,which the Mononucleotides and Dinucleotides core sequence were main types in analysis.3.Based on the statistical analysis of FPKM expression quantity,there were 11 579 genes differentially expressed in ?Wen 138?and 1 217 genes in the ?Zhi Pi? in endocarp developmental periods.There are 12 324 genes differentially expressed in ?Zhipi? and 1 077 genes are differentially expressed in three periods.Alignment the expressed genes in same period from two varieties,2 095 unigenes are expressed in the three periods,199 genes are differentially expressed in three periods.GO functional annotations and KEGG metabolic pathway analysis in differentially expressed genes,it is almost consistent that the function of the differentially expressed genes in each period with metabolism pathways,but exception appeared in specific individual genes and periods.4.Gene functional annotation,metabolism and lignin gene expression analysis in obtained unigenes,168 condition specific genes were detected,and 108 sequence in ?Zhipi? and 60 in ?Wen 138?.Alignment with KEGG database,361 genes associated with lignin biosynthesis,there are 110 genes directly involved in the formation of 11 kinds of key enzyme in lignin synthesis,other 251 genes involved in the whole process of the synthesis of lignin.Comparing key enzyme gene in ?Zhipi? and ?Wen 138? walnut,the expression quantity of PAL genes in ?Zhipi? and other key enzyme genes were higher than in ?Wen 138? on June 13.In three developmental periods,most of the key enzyme genes expression trend is risingdecrease in ?Zhipi? and characterized by increaseing in turn in ?Wen 138? walnut.Walnut endocarp formation was studied by RNA-seq technology and the data were analyzed,the result enriched the application of transcriptome analysis technology in walnut research field.The important references of molecular mechanism of biological characters in walnut endocarp were provided by the RNA-seq technology. |
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