牡丹类psDHN-YSK2基因全长cDNA的克隆与表达分析

为了获得牡丹类脱水素基因的全长cDNA序列,推测其在休眠解除进程中的生物学功能,以不同低温处理时间的牡丹花芽为供试材料,末端快速扩增方法克隆全长cDNA序列,实时定量PCR分析其表达模式。结果表明牡丹类脱水素基因全长cDNA序列为1187 bp,包括831 bp的开放阅读框,114 bp的5’非编码区和242 bp的3’非编码区。其编码的蛋白具有两个植物脱水素蛋白特征性K片段,根据Close的分类方法,属于YSK2类脱水素蛋白。系统发生分析表明psDHN-YSK2基因与葡萄亲缘关系最近。在花芽内休眠解除前期随着低温处理时间的延长psDHN-YSK2表达呈上调趋势。本研究克隆了牡丹psDHN-YSK2的全长cDNA序列,分析了其在花芽内休眠解除过程的表达趋势,暗示了其参与了牡丹花芽的内休眠过程。     

干旱胁迫下小麦叶片脱水素的表达与水分的关系

【目的】研究脱水素对小麦抗旱的作用,阐明脱水素表达与水分的关系。【方法】以2个耐旱性不同的同核异质小麦品系(高抗和低抗)为材料,在盆栽自然干旱胁迫和复水条件下处理小麦幼苗,采用SDS-PAGE和Westernblot方法,分析小麦在干旱和复水条件下叶片的相对含水量、细胞膜相对透性和脱水素表达量的变化。【结果】随土壤干旱胁迫程度的加剧,小麦叶片相对含水量下降,细胞膜相对透性增大,有1条28 ku脱水素特异表达;耐旱性弱的小麦品系叶片中脱水素的表达早于耐旱性强的小麦品系,说明其对干旱胁迫更敏感;而耐旱性强的小麦品系叶片中脱水素蛋白含量高于耐旱性弱的小麦品系;复水后,小麦叶片相对含水量上升,细胞膜相对透性减小,脱水素蛋白在一小段时间内仍存在,说明其表达有一定的延续性。【结论】小麦耐旱性与其受干旱影响的程度和脱水素的表达密切相关。     

玉米脱水素基因家族的鉴定与分析

脱水素(dehydrin,DHN)属于LEA蛋白第二家族成员,是一种植物中广泛存在的亲水性蛋白,在干旱、低温和高盐等非生物胁迫的环境中起到重要作用。通过生物信息学方法对玉米全基因组DHN家族成员进行了鉴定,并进一步对其系统发育、基因结构、染色体定位和基因复制以及表达模式进行了系统分析。结果显示,在玉米DHN基因家族中共有5个家族成员,多物种系统发育进化树分析、基因结构以及基序分析都表明DHN家族成员在进化上具有高度的保守性。基因复制和物种间微共线性分析表明,在5个玉米DHN基因中存在着1对片段复制基因(ZmDHN1-ZmDHN2),玉米、高粱和水稻3个物种间存在2对直系同源基因(ZmDHN2-Sb04g032250.1,ZmDHN2-Os02g44870.1)。通过转录组表达数据分析表明,玉米DHN家族基因在不同发育时期具有不同的组织表达模式;同时,诱导表达模式分析表明ZmDHN基因的表达受到盐和干旱胁迫的显著诱导。该研究结果将为进一步鉴定玉米DHN家族重要的基因成员并对其开展功能分析奠定基础。     

Inheritance of Cold Hardiness and Dehydrin Genes in Diploid Mapping Populations of Blueberry

Summary

Studies of herbaceous plants suggest that cold hardiness is a complex, quantitatively inherited trait. Although development of cold hardiness is an integral part of the life cycle of woody perennial plants, studies on the genetic control of cold hardiness in woody perennials are scarce. A better understanding of the genetic control of cold hardiness would be valuable for developing more effective strategies to increase cold hardiness and, hence, climatic adaptation of woody perennial crops. In blueberry, three major dehydrins of 65, 60, and 14 kDa have been found to increase with cold acclimation and decrease with deac-climation. A comparison of these dehydrin levels among various blueberry cultivars and selections has revealed their level of accumulation to be closely associated with cold hardiness level. Efforts are underway to isolate and map the dehydrin genes of blueberry utilizing blueberry populations that segregate for cold hardiness in order to determine if the dehydrin genes map to or co-segregate with QTLs controlling cold hardiness. Progress has been made toward this goal. Cold hardiness levels were determined for a portion of the blueberry mapping populations (derived from testcrosses of Vaccinium darrowi Camp X V. caesariense Mackenz. F₁s to another V. darrowi and another V. caesariense) using a laboratory controlled freeze-thaw regime, followed by visual assessment of injury to floral buds. As expected, the V. darrowi and V. caesariense parents were found to differ significantly in terms of cold hardiness levels (LT₅₀s of -13°C and -20°C, respectively). Mean cold hardiness level of F₁s (LT₅₀ of -14.7°C) was skewed toward the V. darrowi parents suggesting that cold hardiness is a partially recessive trait. The sequence of a 2.0 kb cDNA clone, which encodes the 60 kDa blueberry dehydrin, was used to map a dehydrin-related gene to current linkage group 12 of the V. caesariense testcross population. A preliminary comparison of the segregation pattern of the dehydrin-related gene to that of the cold hardiness trait suggests that the marker does not segregate with cold hardiness.     

核桃Y2SK2型脱水素基因JrDHN的克隆、表达和单核苷酸多态性分析

采用RT-PCR和RACE技术从‘香玲’核桃（Juglans regia L.‘Xiangling’）叶片中克隆了1个脱水素基因JrDHN（GenBank 登录号KC503061.1）,其全长1 056 bp,具有528 bp的开放阅读框,编码175个氨基酸组成的多肽,具有LEA家族成员的特征多肽序列,属于典型的Y₂SK₂型脱水素。以核桃18S基因为内参,对‘香玲’核桃JrDHN在4 ℃胁迫下的表达模式进行了初步的研究,结果表明低温诱导下叶片中JrDHN的表达增加,4 ℃处理4 h后达到最大值;自然越冬条件下,花芽中JrDHN表达量呈先上升后下降的趋势,在12月份表达量最高,推测JrDHN在核桃对低温胁迫响应中发挥重要功能。‘香玲’JrDHN在雌花中表达量高,其次是叶片和树皮,在花芽中最低。单核苷酸多态性分析JrDHN序列中有30个SNPs位点和11个Indels;单倍型分析显示12份核桃材料可分为10个单倍型,单倍型多样性为0.9697。     

不同类型脱水素在植物低温胁迫应答中的作用

低温是影响植物生长发育和作物产量的非生物胁迫因子之一。植物在长期的进化过程中,通过调节自身的生理及分子变化,形成了对低温胁迫的适应能力。脱水素是一类晚期胚胎丰富蛋白,可保护植物细胞内蛋白质和膜结构等不被低温破坏。不同植物中脱水素的过量表达能够提高植物的低温抗性,植物脱水素的表达和蛋白质的积累与植物耐低温能力密切相关。本文对SKn、Kn、YnSKn、KnS和Yn Kn型脱水素在植物抵御低温胁迫过程中的作用进行综述,旨在为进一步利用脱水素进行植物抗低温分子育种研究提供参考。     

CmRT1, a Ty3-gypsy-like retrotransposon in Castanea mollissima,is associated with a short-catkin mutation

Summary

Retrotransposons are major components of the genomes of most eukaryotic organisms and have resulted in the introduction of desirable traits in many crops, including fruit trees. Here, we describe a Ty3-gypsy-like retrotransposon associated with a short-catkin mutant in Chinese chestnut (Castanea mollissima), resulting in catkins that are < 20% the length of normal staminate catkins. A partial sequence of the retrotransposon, named CmRT1, detected by amplified fragment length polymorphism (AFLP) analysis, and its complete sequence were determined from the genome of Chinese chestnut (Castanea mollissima) using improved Tail-PCR. CmRT1 was 10,067 bp in length and shared high homology in its predicted amino acid sequence and motifs with other Ty3/gypsy-like retrotransposons. The 5’ long terminal repeat (LTR) of CmRT1 contained a TATA box and several cis-elements that were predicted to be important for processes involving abscisic acid, gibberellic acid, and auxins and in stress-mediated responses. Further characterisation of the transposition event that led to the short-catkin phenotype was performed using two pairs of primers that aligned with the flanking region of the LTRs. The expected PCR bands were observed only in genomic DNA from plants that showed the mutation. Finally, cloning and real-time qPCR analysis of an NADP-dependent alkenal double-bond reductase (CmADBR) target gene that was adjacent to CmRT1, revealed that CmADBR expression was significantly down-regulated in the short-catkin mutant. Taken together, these results suggest that the CmRT1 retrotransposon is responsible for the short-catkin phenotype.     

植物脱水素对多种逆境的响应

综述了目前关于脱水素（Dehydrin,DHN）的研究进展,包括脱水素的蛋白结构特征、细胞定位及作用机制,脱水素在生物及非生物逆境下的转基因研究,脱水素的磷酸化修饰,脱水素的结合金属离子、清除活性氧、作为分子伴侣 保护酶活性等特性。展望了利用现代生物技术将脱水素用于提高植物对逆境的耐受力的应用前景,提出今后还需在脱水素作用机制、基因家族功能与生物胁迫相关性等几个方面进行深入研究。     

植物脱水素对多种逆境的响应

综述了目前关于脱水素（Dehydrin，DHN）的研究进展，包括脱水素的蛋白结构特征、细胞定位及作用机制，脱水素在生物及非生物逆境下的转基因研究，脱水素的磷酸化修饰，脱水素的结合金属离子、清除活性氧、作为分子伴侣 保护酶活性等特性。展望了利用现代生物技术将脱水素用于提高植物对逆境的耐受力的应用前景，提出今后还需在脱水素作用机制、基因家族功能与生物胁迫相关性等几个方面进行深入研究。     

东方山羊豆脱水蛋白基因的克隆及初步分析

 采用ＳＭＡＲＴ ＲＡＣＥ 方法,从东方山羊豆盐诱导抑制性差减杂交ｃＤＮＡ 文库中分离到了一个脱水蛋白（GoDHN）基因。该基因ｃＤＮＡ 全长１１６９ｂｐ,开放阅读框８４３ｂｐ,编码２８１ 个氨基酸,编码的蛋白质分子量为２８．７１ｋＤａ。经实时荧光定量ＰＣＲ 分析,GoDHN 基因在东方山羊豆的茎和叶中表达量明显高于根中表达量,并且基因表达受ＡＢＡ、ＮａＣｌ和ＰＥＧ 的诱导,随着诱导时间的增加,表达量呈持续增长趋势。这些结果表明,DHN 基因在东方山羊豆的抗逆性中可能起到重要的调控作用。本研究成功构建了ｐＣＡＭＢＩＡ１３０２-ＤＨＮ 植物表达载体,为下一步转基因研究奠定了基础。