CloneIRD:面向代码溯源的克隆代码继承关系判定方法

随着开源软件的广泛使用,代码溯源成为管理软件源代码、降低潜在风险的重要技术手段。基于代码克隆检测的大规模代码溯源分析,从其检测结果中鉴别代码克隆对之间的继承关系,对代码来源追踪、组件依赖关系分析、软件脆弱性分析以及代码缺陷修复等具有重要意义。目前,已有方法在原始代码片段存在微小修改的情况下,会产生许多误判,并且检测克隆对的效率也有待提高。针对上述问题,提出了代码溯源中克隆代码继承关系的判定方法CloneIRD,包括一个基于自研快速分布式克隆检测工具FastDCF的代码溯源分析框架,以及该框架的核心算法——基于代码演化信息的克隆代码继承关系判定算法EIHR。为验证框架和算法的有效性,首先设计并实现了CloneIRD方法,并在Linux内核V4.9和V4.12的开源代码上进行了实验。实验结果表明,CloneIRD方法能够有效判定代码溯源结果中克隆对的继承关系,且基于FastDCF的溯源分析框架能够胜任大规模代码的溯源分析任务。

Gene Cloning and Bioinformatics Analysis of phoR Gene from Vibrio alginolyticus HY9901

PhoR is a histidine kinase in a two-component regulatory system that regulates phosphorus metabolic pathways and undertakes the key mission of information transmission in pathogenic bacteria.The full-length phoR gene was successfully cloned from the Vibrio alginolyticus HY9901 strain.A comprehensive analysis of the cloned gene was conducted using bioinformatics.Sequence analysis revealed that the total length of the phoR gene(GenBank accession No.:KJ958404.1)is 1299 bp,with the coding region containing a total of 432 amino acid residues.The phylogenetic tree of PhoR revealed that it belongs to the same subclade as V.diabolicus.The SMART program was employed for the purpose of functional domain prediction,which revealed that PhoR possesses three major functional domains:PAS(amino acids 98-166),HisKA(amino acids 205-272),and HATPase_c(amino acids 317-429).

遵循解剖特点的A-T皮瓣在面部软组织缺损修复中的应用

目的:探讨遵循解剖特点的A-T皮瓣在面部软组织缺损修复中的应用效果。方法:笔者对22例面部软组织缺损患者行A-T皮瓣修复,根据患者各个区域软组织缺损情况、周边相连组织特点、解剖纹理及langer线走向综合设计遵循解剖特点的A瓣的底边。唇周、鼻翼缘设计在皮肤黏膜交界区;眼周设计线多位于睑缘;眉区设计线多位于眉上或者眉下眉毛和皮肤交界处;睑袋区设计线可在泪沟线;额部近发际线可设计在发际线。根据各区解剖特点,皮瓣切口线可呈弧形、对应线长短不一定相等。将外伤创面或者需切除物设计包含在A瓣内,根据创面或者需切除物大小,沿轮廓线或自然交界线向两侧延长底边至合适长度。沿设计线切开皮肤和皮下组织,掀开皮瓣,对合A瓣底边两点并缝合成T瓣,各对应边对位缝合。结果:22例患者A-T皮瓣修复术后,术区未见明显缺血、瘀血和皮瓣收缩,皮瓣全部存活,切口瘢痕形成不明显。结论:遵循解剖特点的A-T皮瓣在修复面部相关区域软组织缺损时效果较好,对周边形态影响小,瘢痕形成少,值得临床推广应用。

机器学习法预测不同应力比6005A-T6铝合金疲劳裂纹扩展速率

本文开展了不同应力比(R为0.05,0.1,0.3,0.5,0.6)条件下的6005A-T6铝合金紧凑拉伸试样疲劳裂纹扩展速率试验,在此基础上采用BPNN、SVR、KNN和XGboost共4种机器学习方法,比较了4种机器学习方法预测6005A-T6铝合金疲劳裂纹扩展速率的能力;作为对照组,利用试验数据拟合得到了传统的Forman疲劳裂纹扩展率方程。结果表明:应力比R对6005A-T6铝合金裂纹扩展速率存在显著影响,相同应力强度因子范围ΔK下,裂纹扩展速率da/d N随R增大而提高,且R的增大会使裂纹更早进入稳定扩展阶段;4种机器学习模型建立的裂纹扩展率预测内推模型均能体现裂纹扩展过程中速率的非线性变化特征,且对试验数据的拟合系数r^(2)均达0.99以上,普遍高于Forman方程的0.82;4种机器学习模型建立的裂纹扩展率预测外推模型,BPNN算法的拟合系数r^(2)仍高于0.99;KNN算法消耗的训练时间最短,XGboost算法具有最佳的疲劳裂纹扩展速率内推预测能力,而BPNN算法则具有更好的扩展率外推预测性能表现。

Economical phase-covariant cloning with multiclones

This paper presents a very simple method to derive the explicit transformations of the optimal economical 1 to M phase-covariant cloning. The fidelity of clones reaches the theoretic bound [D＇Ar]ano G M and Macchiavello C 2003 Phys. Rev. A 67 042306]. The derived transformations cover the previous contributions [Delgado Y, Lamata Let al, 2007 Phys. Rev. Lett. 98 150502] in which M must be odd.

Superovulation of the Cloned Cattle Derived from Somatic Cells and the Transfer of the Vitrified-Thawed Embryos of the Cloning Cattle

In this experiment, it was designed to carry out superovulation on the two cloned cattles, vitrification and transfer of the embryos recovered from them. First of all, it was carried out vitrification on embryos obtained by IVF. Results showed that there were no significant differences between the blastocysts (obtained by IVF) vitrified in EPS10 and these in EPS20 on the resuscitative rate and the developmental rate. The hatched rate of the blastocysts vitrified in EPS10 (31.3%, 35/112) was significantly higher than that in EPS20 (12.2%, 13/107)(P<0.01), so EPS20 was selected as the vitrification solution to freeze the embryos recovered from the cloned cattle. After superovulation, six (four usable embryos) and ten (nine usable embryos) embryos were respectively recovered from Kangkang and Shuanghuang. Two embryos were selected from the recovered embryos of each cloned cattle to freeze in EPS20, subsequently thawed and transferred into luteal ipsilateral uterine horns of 4 Holstein recipient cows after synchronization of estrus, respectively. At last, one recipient cow (No. 9908) became pregnant and delivered one healthy calf (descendant of the cloned cattle-Shuangshuang). The results of this experi- ment show that the cloned cattle as well as common cattle had better response to the exotic FSH and better ability to multiovulation, the embryos recovered from the cloned cattle can be vitrificated.

An integrated microfluidics platform with high-throughput single-cell cloning array and concentration gradient generator for efficient cancer drug effect screening

Background:Tumor cell heterogeneity mediated drug resistance has been recognized as the stumbling block of cancer treatment.Elucidating the cytotoxicity of anticancer drugs at single-cell level in a high-throughput way is thus of great value for developing precision therapy.However,current techniques suffer from limitations in dynamically characterizing the responses of thousands of single cells or cell clones presented to multiple drug conditions.Methods:We developed a new microfluidics-based“SMART”platform that is Simple to operate,able to generate a Massive single-cell array and Multiplex drug concentrations,capable of keeping cells Alive,Retainable and Trackable in the microchambers.These features are achieved by integrating a Microfluidic chamber Array(4320 units)and a sixConcentration gradient generator(MAC),which enables highly efficient analysis of leukemia drug effects on single cells and cell clones in a high-throughput way.Results:A simple procedure produces 6 on-chip drug gradients to treat more than 3000 single cells or single-cell derived clones and thus allows an efficient and precise analysis of cell heterogeneity.The statistic results reveal that Imatinib(Ima)and Resveratrol(Res)combination treatment on single cells or clones is much more efficient than Ima or Res single drug treatment,indicated by the markedly reduced half maximal inhibitory concentration(IC50).Additionally,single-cell derived clones demonstrate a higher IC_(50) in each drug treatment compared to single cells.Moreover,primary cells isolated from two leukemia patients are also found with apparent heterogeneity upon drug treatment on MAC.Conclusions:This microfluidics-based“SMART”platform allows high-throughput single-cell capture and culture,dynamic drug-gradient treatment and cell response monitoring,which represents a new approach to efficiently investigate anticancer drug effects and should benefit drug discovery for leukemia and other cancers.

Cloning of PsMYB62 and analysis of cadmium resistant in Potentilla sericea

Potentilla sericea is a heavy metal hyperaccumulator landscaping plant.MYB transcription factors play an important role in regulating plant stress response to adversity.However,there are few studies on MYB transcription factors in stress tolerance in Potentilla sericea.In this study,the PsMYB62 gene was successfully cloned from Potentilla sericea.Methods:Bioinformatic analysis and real-time quantitative PCR(qPCR)methods were used to evaluate this gene.The transgenic A.thaliana were obtained by flower dipping and the gene function was identified by determining physiological indicators under cadmium stress.Results:The open reading frame of PsMYB62 is 942 bp,which encodes 313 amino acids(aa)and belongs to the R2R3 MYB transcription factor.The plant overexpression vector PBI121-PsMYB62-GFP was constructed and successfully transferred into A.thaliana.The relative expression level of PsMYB62 was significantly increased by CdCl_(2),NaCl,ABA,and mannitol treatments.The germination rate of transgenic seeds was higher than those of wild type(WT)and empty vector(EV)under different concentrations of cadmium treatment.Upon treatment with 100μmol·L^(−1)of CdCl_(2)·2.5H_(2)O,the activities of superoxide dismutase(SOD),peroxidase(POD),and catalase(CAT)in the transgenic plants were significantly higher than those in the WT and EV.The contents of H_(2)O_(2),O_(2)·−and malondialdehyde(MDA)in transgenic lines were increased,but lower than those in WT and EV.The expression levels of AtGSH,AtPCS,and AtNAS4 that were related to the regulation of cadmium were increased,but the expression levels of transgenic lines were higher than those of WT and EV.Conclusion:The above results showed that PsMYB62 could be induced by cadmium and could improve the cadmium resistance of plants.

Cloning and Functional Validation of Mung Bean VrPR Gene

For the purpose of functional validation,the mung bean(Vigna radiata)VrPR gene was cloned and overexpressed in Arabidopsis thaliana.Thefindings revealed that the ORF of VrPR contained 1200 bp,in which 399 amino acids were encoded.Bioinformatics analysis showed that the VrPR protein belonged to the NADB Rossmann superfamily,which was one of the non-transmembrane hydrophilic proteins.VrPR was assumed to have 44 amino acid phosphorylation sites and be contained in chloroplasts.The VrPR secondary structure comprised of random coil,αhelix,βangle,and extended chain,all of which were quite compatible with the anticipated tertiary structure.Moreover,analysis of the phylogenetic tree indicated that the soybean PR(Glyma.12G222200)and VrPR were closely related.Furthermore,chlorophyll content in leaves is markedly increased in Arabidopsis when VrPR is overexpressed.Ourfindings will serve as a reference for more functional studies on the PR genes in mung bean.

Cloning and Bioinformatics Analysis of vscB Gene of T3SS Chaperone of Vibrio alginolyticus

[Objectives]To clone and analyze the vscB gene of Vibrio alginolyticus HY9901 by bioinformatics.[Methods]A pair of specific primers were designed according to the vscB gene sequence of Vibrio alginolyticus HY9901.The full length of the primers was cloned by PCR and analyzed by bioinformatics.[Results]The vscB gene was 429 bp long,encoding 142 amino acids,with a theoretical molecular weight of 16.4 kDa and a pI value of 5.48.Amino acid sequence analysis of VscB showed that VscB was not a secretory protein,without signal peptide and transmembrane region,and there were protein kinase C phosphorylation site and casein kinase II phosphorylation site in the sequence.Homologous comparison of amino acid sequences showed that VscB of V.alginolyticus had the highest protein similarity with Vibrio Parahaemolyticus,reaching 91%.Phylogenetic tree analysis showed that the corresponding proteins of V.alginolyticus VscB,Vibrio Parahaemolyticus and Vibrio diabolicus were clustered in the same subfamily.Functional domain analysis showed that it had CesT family domain.Tertiary structure prediction showed that there were 3α-helices and 5β-turns in VscB protein.[Conclusions]This study provided a theoretical basis for further study on the function of chaperone of V.alginolyticus.

The Clones’Struggle for Happiness under the Doomed Fate in Never Let Me Go

Japanese British writer Kazuo Ishiguro is one of the leading writers in contemporary British literature.He has always been committed to creating works with universal significance.Responsibility and destiny are themes that run through his works.His novel Never Let Me Go tells a story of a group of clones growing up in the Hailsham,who are given the mission to donate organs at birth.So,there is no doubt that they will inevitably end their lives in the process of donating organs to human beings again and again.The tragic life of clones is determined by the motivation of human to create them.

An Analysis on the Role Identification of Clones in Never Let Me Go

With the guidance of Erikson’s identity theory,the article analyzes the clones’identity exploration through role identification in Never Let Me Go.It interprets the clones’puzzlement about their identity,and the process of their identity quest as well as role identification.Through the specific analysis,it is concluded that the clones,represented by Kathy,Tommy and Ruth,have gained self-certainty and social identity by realizing the role identity as a“carer”and a“donor”,in the meantime,they have constructed their identity as social persons with souls like ordinary people.Furthermore,the findings shows that Never Let Me Go is actually a microcosm of human’s quest for identity.Ishiguro aims to express his meditation on human life in the novel:human life is a process of seeking self-identity and social roles,and then fulfilling the obligations of the roles,which confirms Ishiguro’s internationalism that he attempts to convey his contemplation on human existence through his works.

羽衣甘蓝BoLMI基因的克隆及时空表达分析

【目的】探究羽衣甘蓝裂叶表型形成的分子机制,研究BoLMI基因对裂叶形成的作用。【方法】以裂叶羽衣甘蓝DH系为试验材料,利用同源克隆技术,成功获取基因完整的cDNA序列,将其命名为BoLMI。【结果】序列分析发现,BoLMI基因全长531个碱基对,共编码176个氨基酸,其蛋白分子量为20 789.35 Da,理论等电点为7.24。根据Pfam保守结构域分析,BoLMI蛋白包含homeobox保守结构域。系统发育分析结果显示,羽衣甘蓝的BoLMI基因与甘蓝型油菜以及结球甘蓝的BoLMI基因同属于一个分支,亲缘关系较近。BoLMI蛋白是位于细胞核内的可溶性蛋白,包含1个跨膜结构域,无信号肽序列。qRT-PCR分析表明,BoLMI基因在裂叶羽衣甘蓝的幼苗期表达水平较高,在莲座期表达水平较低;在羽衣甘蓝莲座期,裂叶品种不同部位叶片BoLMI基因的表达水平均高于圆叶品种,其中在第1片叶的表达量最高。裂叶品种和圆叶品种不同叶片BoLMI基因的表达量差异显著。【结论】推测BoLMI基因在羽衣甘蓝裂叶形成中起重要作用,为加速羽衣甘蓝叶形育种提供了理论基础。

洋葱AcSCL4的克隆及其功能分析

【目的】初步探讨AcSCL4在洋葱成花中的功能,为解析洋葱成花分子调控机制提供理论基础。【方法】以长日生态型洋葱品系SB2为试验材料,根据已有的洋葱转录组数据,通过qRT-PCR筛选目的基因AcSCL4后进行基因克隆,并对其进行生物信息学分析;构建AcSCL4基因的过表达载体pB221-3300-AcSCL4并转化农杆菌,采用花序浸染法浸染拟南芥,并对T_(3)代纯系转基因植株进行表型分析和开花相关基因表达量分析。【结果】洋葱AcSCL4基因CDS长1515 bp,编码504个氨基酸;AcSCL4蛋白N端高度变异,C端有GRAS结构域。聚类分析发现,AcSCL4与石刁柏和棉花的SCL4蛋白亲缘关系较近。与野生型拟南芥相比,AcSCL4过表达植株表现晚花表型。荧光定量PCR结果表明,与野生型拟南芥相比,AcSCL4过表达植株开花调控基因CO和FT的表达量极显著下降。【结论】AcSCL4通过调控CO及FT基因表达来抑制洋葱开花。

甘薯U6启动子克隆及其转录活性分析

在CRISPR/Cas9基因编辑系统中,U6启动子可以通过驱动sgRNA的表达来编辑目的基因,因而其转录活性会影响基因编辑效率。尽管人们已经在拟南芥、玉米、大豆、棉花等植物中开展了U6启动子的克隆与应用研究,然而目前在甘薯中U6启动子的相关研究还未见报道。本研究利用拟南芥的AtU6 SnRNA的保守序列在三浅裂野牵牛(Ipomoea trifida)基因组数据库中搜索候选U6 RNA,然后在其上游搜索到2个不同的候选U6启动子,长度分别为526 bp(IbU6p-1)和532 bp(IbU6p-2);序列比对分析结果显示,甘薯的U6启动子具有上游序列元件(USE)以及TATA-box,其序列也与拟南芥高度相似。然后,利用获得的U6启动子核酸序列构建了能够驱动萤火虫荧光素酶基因(LUC)表达的重组框U6::LUC。最后,将含有上述重组载体的根癌农杆菌瞬时转化到本氏烟草叶片和甘薯愈伤组织中,并通过荧光成像技术分析荧光素酶活性。结果发现,在烟草及甘薯愈伤组织中2个甘薯U6启动子均能驱动LUC基因表达,具有转录活性。同时,IbU6p-2的转录活性无论是在烟草叶片中还是在甘薯愈伤组织中都显著高于拟南芥U6启动子。本研究结果为进一步发展甘薯基因编辑技术提供了参考。

草莓FaWRKY70基因克隆与表达模式分析

【目的】WRKY是植物特异性转录因子家族之一,参与植物多种生命活动,但在草莓中鲜见WRKY70相关报道。深入解析WRKY70同源基因在草莓响应胁迫过程中的作用,加快分子育种技术应用,培育新草莓种质资源。【方法】用同源克隆法从‘红颜’草莓果实中克隆得到FaWRKY70基因,用生物信息学分析其保守结构域、理化性质、蛋白质结构及进化关系等,并结合qRT-PCR数据进行表达模式分析。【结果】FaWRKY70基因全长1020 bp,编码339个氨基酸;同源基因比对发现,FaWRKY70与苹果、牡丹等同科物种氨基酸序列相似度较高,且同源基因多与植物对生物、非生物胁迫响应相关,暗示FaWRKY70可能参与草莓抵御胁迫的过程;FaWRKY70在草莓不同器官中均有表达,且差异显著,在花中表达量最高,在果实中最低;水杨酸处理后,FaWRKY70基因快速响应,表达量在3 h后达到最高,随后逐渐降低;茉莉酸甲酯处理后FaWRKY70基因受诱导响应程度小,整体呈下调趋势。【结论】FaWRKY70能通过不同响应方式参与草莓生命活动及激素信号转导过程。

康乃馨抗尖孢镰刀菌无性系诱变技术

枯萎病是康乃馨鲜切花种植过程中较为严重的真菌病害之一,该病的病原菌是尖孢镰刀菌康乃馨专化型(Fusarium oxysporum f.sp.dianthi),培育和合理种植抗病品种是防控康乃馨枯萎病最有效的方法之一。应用植物体细胞无性系变异及真菌毒素加压筛选技术,可以定向培育康乃馨抗病育种材料并加快抗病品种选育速度,为康乃馨抗病育种提供新的思路。为获得抗枯萎病的康乃馨育种中间材料,以感枯萎病的康乃馨多花品种紫蝴蝶组培苗为试验材料,诱导愈伤组织并进行悬浮培养建立悬浮培养系,再用甲基磺酸乙酯(EMS)诱变后添加尖孢镰刀菌毒素粗提液筛选抗病细胞系。结果表明,诱导愈伤组织最适宜的培养基是Murashig-Skoog培养基+麦草畏1.0 mg/L;筛选出EMS最佳处理组合为0.4%处理4 h;在80.0%的粗毒素培养基上培养10 d是康乃馨抗尖孢镰刀菌无性系筛选较适宜的选择压;诱导康乃馨再生植株较好的激素组合是苄氨基腺嘌呤(BA)0.5 mg/L+噻苯隆(TDZ) 0.1 mg/L+萘乙酸(NAA)0.1 mg/L;经人工接种尖孢镰刀菌进行抗病性鉴定后发现,紫蝴蝶抗病无性系的病情指数为45,为中抗水平。

杨树无性系表型性状及ISSR分子标记遗传多样性分析

【目的】综合表型性状及分子标记多样性分析探明供试的62个杨树无性系的遗传多样性,为杨树进一步遗传改良奠定基础。【方法】选取地径、苗高、叶面积、叶长、叶宽、叶柄长、叶绿素、侧枝数、叶厚、单株总叶片、叶片干质量、叶片含水率等12个表型性状和ISSR分子标记对杨树无性系个体间进行遗传多样性分析,利用PopGen 32、SPSS 16.0及NTsys 2.10e等软件分别计算多样性指数、进行表型性状间的方差分析以及对各无性系进行UPGMA法聚类分析。【结果】侧枝数、单株叶片数、叶片干质量、叶面积以及地径的变异系数均达到了10%以上,叶柄长、叶长、苗高的变异系数也达到了8%以上的水平。利用5条ISSR引物检测到62份杨树无性系多态性谱带百分率平均为89.04%,基因多样度平均为0.4074,Shannon多样性指数(I)平均为0.5304。采用UPGMA法构建的形态和分子标记聚类图将供试材料聚成的类群均有家系内聚为一类的趋势,但也存在较大的差异;表型性状聚类分析主要是根据叶片的相关性状相似度越高的被聚为一个类群;分子标记聚类主要呈现出亲缘关系越近的无性系越容易聚为一个类群的聚类规律。【结论】供试的杨树无性系间表型性状分化程度高,具有丰富的遗传多样性,研究结果为杨树种质资源的改良、种质创新及多元化开发利用提供科学依据。

番木瓜环斑病毒甜瓜分离物的基因组及其侵染性克隆

番木瓜环斑病毒(papaya ringspot virus,PRSV)是瓜类作物主要病毒之一,分析了甜瓜分离物HaNHK10的基因组序列和分子变异,构建了具有侵染性的全长cDNA克隆。结果显示,HaNHK10分离物基因组全长为10332 nt,与其他分离物的核苷酸和氨基酸序列一致性分别为74.60%~97.80%和85.30%~98.50%。基于全基因组序列的系统进化分析显示,HaNHK10与来自中国的所有分离物均聚集于II组中,并与中国山东的西葫芦分离物PRSV-SD亲缘关系最近。接种试验显示,HaNHK10分离物的全长cDNA克隆具有侵染性,它能系统侵染甜瓜、黄瓜、西瓜、南瓜、西葫芦和瓠瓜6种作物,经接种产生的病毒后代也能够通过摩擦接种侵染植株。

大肠杆菌多酚氧化酶的分子克隆及异源高效表达

菜籽粕、棉籽粕等非粮饲料资源存在的酚类化合物芥子碱、棉酚等抗营养因子,严重影响了其饲喂价值。本课题组筛选出的一株大肠杆菌属芥子碱降解菌SDB2已被证实能通过分泌多酚氧化酶来降解芥子碱和棉酚,本试验旨在运用基因工程技术克隆SDB2多酚氧化酶基因,实现其高效表达,为SDB2多酚氧化酶在饲料工业上的规模化应用奠定基础。从大肠杆菌SDB2中克隆出多酚氧化酶基因,将其与质粒pET28a连接后转化至大肠杆菌BL21感受态细胞中培养,通过PCR鉴定及质粒酶切验证方法构建重组菌株,同时对重组克隆基因进行生物信息学分析。采用“2×2”交叉试验设计,以温度和诱导剂IPTG浓度两个影响因素诱导SDB2多酚氧化酶基因在重组菌株中高效表达,经SDS-PAGE和WB双重验证后,确立最佳诱导条件,最终纯化出目标酶蛋白。结果表明:(1)成功克隆SDB2多酚氧化酶基因,构建出异源表达重组菌株。(2)克隆出的SDB2重组多酚氧化酶基因含目标核苷酸741个,总编码氨基酸263个(含标签氨基酸20个),氨基酸组成的蛋白相对分子质量为28499.0,理论等电点为6.94,据此预测出SDB2重组多酚氧化酶三维结构模型。(3)确立SDB2多酚氧化酶异源高效表达的最佳诱导条件为温度37℃,IPTG浓度1 mmol,该条件下表达出大量可溶性酶蛋白,有利于工业化生产。本试验条件下,大肠杆菌SDB2多酚氧化酶成功实现了分子克隆及异源高效表达,为我国非粮饲料资源实现高效利用提供了技术参考。