目的 對(duì)白蠟樹(shù)Fraxinus chinensis WRKY基因家族進(jìn)行鑒定、生物信息學(xué)和密碼子偏好性分析。方法 基于白蠟樹(shù)全長(zhǎng)轉(zhuǎn)錄組數(shù)據(jù)，利用生物信息學(xué)方法對(duì)其WRKY基因家族進(jìn)行鑒定，并對(duì)其理化性質(zhì)、系統(tǒng)發(fā)育、保守基序進(jìn)行分析；基于白蠟樹(shù)葉片和枝皮的二代轉(zhuǎn)錄組數(shù)據(jù)，對(duì)白蠟樹(shù)WRKY基因家族的表達(dá)模式進(jìn)行分析，并采用實(shí)時(shí)熒光定量PCR方法進(jìn)行驗(yàn)證；利用CodonW、EMBOSS等程序?qū)γ艽a子偏好性進(jìn)行分析，并進(jìn)行最優(yōu)密碼子分析、中性繪圖、ENC-plot和PR2偏倚分析。結(jié)果 共鑒定得到49個(gè)WRKY轉(zhuǎn)錄因子，均為酸性、親水蛋白，編碼氨基酸數(shù)目為426～2 175，相對(duì)分子質(zhì)量33 933.48～179 137.94，理論等電點(diǎn)4.91～5.27，不穩(wěn)定蛋白占比65.31%，預(yù)測(cè)均定位到細(xì)胞核；系統(tǒng)進(jìn)化表明FcWRKY基因家族成員歸為Ⅰ（11個(gè)）、Ⅱ（30個(gè)）、Ⅲ（7個(gè)）大類以及2個(gè)特殊成員（FcWRKY23和FcWRKY37），其中第Ⅱ類分為Ⅱa（5個(gè)）、Ⅱb（5個(gè)）、Ⅱc（8個(gè)）、Ⅱd（7個(gè)）、Ⅱe（7個(gè)）5個(gè)亞類；Motif 1和Motif 2存在于所有的WRKY轉(zhuǎn)錄因子中，是其保守序列；二代轉(zhuǎn)錄組數(shù)據(jù)篩選到16個(gè)WRKY基因在葉片和枝皮中具有顯著差異表達(dá)，其中僅有FcWRKY11在枝皮中的表達(dá)量高于葉片。白蠟樹(shù)FcWRKYs偏好使用以A/U結(jié)尾的密碼子，并且偏好性較弱；確定了3個(gè)最優(yōu)密碼子，分別是CUU、AGG、和UAU；中性繪圖分析和ENC-plot分析均表明自然選擇是影響其密碼子偏好性的主要因素，PR2偏倚分析結(jié)果表明突變壓力亦影響其密碼子偏好性。結(jié)論 鑒定了白蠟樹(shù)WRKY基因，其中8個(gè)WRKY基因表達(dá)量與轉(zhuǎn)錄組學(xué)測(cè)序結(jié)果趨勢(shì)一致，為研究白蠟樹(shù)WRKY的基因功能奠定基礎(chǔ)。

Objective The aim of this study was to perform identification and bioinformatics and codon preference analysis of WRKY gene family of Fraxinus chinensis. Methods Based on the full-length transcriptome data of F. chinensis, the WRKY gene family was identified using bioinformatics methods and their physicochemical properties, phylogeny, conservative motifs were analyzed. Based on the second generation transcriptome data, expression patterns of WRKY gene family were analyzed and verified by real time fluorescence quantitative PCR. The codon bias was analyzed by CodonW and EMBOSS program, the optimal codon analysis, neutral plotting, ENC plot and PR2 bias analysis were also performed. Results A total of 49 WRKY genes were identified, all of them were acidic and hydrophilic proteins with an amino acid sequence varying from 426 to 2 175, molecular weight between 33 933.48 and 179 137.94, an isoelectric point between 4.91 and 5.27, unstable proteins accounted for 65.31%, and all of them were located in the nucleus. The phylogenetic tree showed that FcWRKY genes were assigned to Ⅰ (11),Ⅱ (30), Ⅲ (7) three categories and two special members (FcWRKY23 and FcWRKY37), the second category was divided into five subclass, including Ⅱa (5), Ⅱb (5), Ⅱc (8), Ⅱd (7) and Ⅱe (7); Conserved motif analysis showed that Motif 1 and Motif 2 were found in all WRKY transcriptional factors. A total of 16 significant differential expression WRKY genes in leaves and bark were identified, of which only FcWRKY11 was high expressed in bark than in leaves. The results of codon bias analysis showed that the FcWRKYs prefered to use codons ending in A/U weak codon usage. The optimal codons were identified, which were CUU, AGG and UAU; The results of neutrality plot analysis and ENC plot analysis indicated that natural selection was the main factor affecting its codon bias, PR2 bias analysis indicated that mutation pressure also had impact on codon preference. Conclusion It laid a theoretical foundation for the study of WRKY gene function in F. chinensis.

R286.12

陜西中醫(yī)藥大學(xué)秦藥特色資源研究開(kāi)發(fā)重點(diǎn)實(shí)驗(yàn)室開(kāi)放課題（KF202326）；陜西省教育廳青年創(chuàng)新團(tuán)隊(duì)建設(shè)科研計(jì)劃項(xiàng)目（21JP031）；陜西高校青年創(chuàng)新團(tuán)隊(duì)資助項(xiàng)目（陜教［2019］90號(hào)）；陜西中醫(yī)藥大學(xué)校級(jí)科研課題（2023GP32）；云南省科技廳基礎(chǔ)研究專項(xiàng)資助項(xiàng)目（202101AU070005）