| Low temperature is one of the main abiotic stresses that limit the growth and development of plants.It is of great significance to study the mechanism of forest response to low temperature chilling injury to improve the cold resistance of trees and expand the forest area.In this study,the seedlings of birch were treated at low temperature,and the molecular mechanism of birch response to low temperature stress was studied by transcriptome sequencing analysis.All AP2/ERF transcription factors in birch genome were identified and analyzed.The main results are as follows:1.The RNA of leaves of white birch treated at 6℃ for 0 h,1 h,1.5 h,2 h,2.5 h and 3 h were sequenced by Illumina Hiseq 2000 sequencing platform and 3,197(1,656 up-regulated and 1,541 down-regulated),881(494 up-regulated and 387 down-regulated),79(67 up-regulated and 12 down-regulated)and 859(470 up-regulated)were obtained respectively.There were 389 down-regulated genes and 451 differentially expressed genes(378 up-regulated and 73 down-regulated).2.Through GO enrichment analysis of differential genes,3491 differential genes were annotated into 1,002 GO items under 3 classifications.Among them,there are more genes enriched in cell process,metabolic process and stimulus response items in biological process classification,and more genes are enriched in organelles and cell components in cell component classification.there are more genes enriched in items such as catalytic activity and adhesion in molecular function classification.3.A total of 119 DEGs were identified to be involved in Ca2+signal transduction(50)and plant hormone signal transduction(69),205 DEGs were involved in starch and sucrose(30),lipid(109),amino acid(40)synthesis and metabolism,phenylpropanoid biosynthesis pathway(26)and other secondary metabolic processes,20 DEGs were involved in photosynthesis.362 transcription factors are distributed in 58 transcription factor families,including AP2/ERF,C2H2,MYB-HB-like,WRKY,bHLH,WD40-like and GRAS families.4.Gene Bpev01.c0480.g0081(calmodulin-like CML38),Bpev01.c1074.g0005(calmodulin-like CML25),Bpev01.c1074.g0001(Calcium-binding EF-hand family protein),Bpev01.c2029.g0005(calmodulin-like protein),Bpev01.c0154.g0008(POD),Bpev01.c0015.g0143(N-acetyl-l-glutamate synthase)and Bpev01.c0148.g0010(branched chain amino acid transferase)were up-regulated at a high level under low temperature stress.13 members of AP2/ERF transcription factor family were continuously upregulated.5.The expression patterns of 12 DEGs genes were verified by real-time fluorescence quantitative PCR(qRT-PCR),and the correlation analysis was carried out with the results of transcriptome.The results showed that the expression of qRT-PCR was consistent with that of transcriptome.6.The basic information of 146members of the AP2/ERF family of birch was determined,including open reading frame length,isoelectric point,molecular weight,number of introns and exons,etc.By analyzing the characteristics of their conserved domains and gene structure,the members of the AP2/ERF family of birch were divided into five different subfamilies,AP2(23),ERF(57),DREB(62),RAV(3)and Soloist(1).7.The members of AP2/ERF gene family were located on 14 chromosomes of birch,but the distribution was not uniform.The number of distributions on chromosome 12 was the least,and that on chromosome 14 was the most.In the AP2/ERF gene family of birch,a total of 25.47%(27/106)of genes were replicated,including 5 pairs of fragment repeat genes and 22 pairs of tandem repeat genes.8.Through the analysis of RNA-seq data,it was found that 42 AP2/ERF genes of birch were differentially expressed under low temperature stress.The expression of BpAP2ERF65(CBF4),BpAP2ERF73(CBF4),BpAP2ERF117(DDF1),BpAP2ERF120(CBF4),BpAP2ERF122(CBF2),BpAP2ERF127(CBF3),BpAP2ERF128(CBF2)of DREB-A1 subfamily,BpAP2ERF8(ERF5),BpAP2ERF9(ERF5),BpAP2ERF11(ERF5),BpAP2ERF14(ERF5),BpAP2ERF15(ERF5),BpAP2ERF47(ERF105)of ERF-B3 subfamily and BpAP2ERF85(RRTF1)of ERF-B4 subfamily were significantly up-regulated.BpAP2ERF44,BpAP2ERF74,BpAP2ERF90 and BpAP2ERF144 of AP2 subfamily were down-regulated. |
