生物多样性 ›› 2018, Vol. 26 ›› Issue (5): 445-456.doi: 10.17520/biods.2018058

• 综述 • 上一篇    下一篇

传粉网络构建的定性定量分子研究: 应用与展望

郎丹丹1, 2, 唐敏1, 3, 周欣1, 3, *()   

  1. 1 中国农业大学北京食品营养与人类健康高精尖创新中心, 北京 100193
    2 中国农业大学食品科学与营养工程学院, 北京 100083
    3 中国农业大学植物保护学院, 北京 100193
  • 收稿日期:2018-02-15 接受日期:2018-05-11 出版日期:2018-05-20
  • 通讯作者: 周欣 E-mail:xinzhou@cau.edu.cn
  • 作者简介:

    # 共同第一作者

  • 基金项目:
    国家自然科学基金(31772493)

Qualitative and quantitative molecular construction of plant-pollinator network: Application and prospective

Dandan Lang1, 2, Min Tang1, 3, Xin Zhou1, 3, *()   

  1. 1 Beijing Advanced Innovation Center for Food Nutrition and Human Health, China Agricultural University, Beijing 100193
    2 College of Food Science and Nutritional Engineering, China Agricultural University, Beijing 100083
    3 College of Plant Protection, China Agricultural University, Beijing 100193
  • Received:2018-02-15 Accepted:2018-05-11 Online:2018-05-20
  • Contact: Zhou Xin E-mail:xinzhou@cau.edu.cn
  • About author:

    # Co-first authors

传粉者是重要的生态功能提供者, 在维持稳定的生态系统和高效的农业生产力中发挥着重要作用。因此, 传粉网络的构建和监测工作对评价生态系统平衡和调控农业生产至关重要。该工作的基础就是通过对传粉者及植物的物种鉴定构建其相关性。传统的形态学物种鉴定对分类学专家的专业知识、时间和经验都提出较高的要求。DNA条形码和高通量测序技术(high-throughput sequencing, HTS)的发展及其在传粉网络研究中的应用, 提供了高效、准确鉴定传粉者与植物的方法, 大大提高了传粉网络构建的效率。本文阐述了传粉网络研究相关的研究方法和技术进展, 并提出利用高通量测序技术结合无PCR扩增(PCR-free)的“超级条形码”技术, 有望实现以更高的灵敏度和分辨率对混合物种样品进行定性及相对定量的监测。该方法的有效性已在其他生物多样性研究中得以验证, 在传粉网络研究中虽处于初始阶段, 但应用前景广阔。

关键词: 线粒体, 叶绿体, 花粉, 宏条形码, 宏基因组, PCR-free, 定量

Pollinators serve key ecological functions, ensuring stable ecosystems and high agricultural yields. Hence, assessing ecosystem health and effects of agricultural management would benefit from understanding and monitoring pollination networks, which involves identifications of pollinators and pollinated plants. Classic approaches of morphology-based identification of plants and pollinators can be time-consuming, labor-intensive and costly, and require highly specialized taxonomic expertise. In comparison, DNA barcoding and high-throughput sequencing technologies can provide efficient and accurate identifications of plants and their pollinators, which may facilitate construction of pollination networks. Here we propose using sequencing technologies with a PCR-free genome-skimming work frame, using "super DNA barcode" as a new method to assess plant-pollinator networks. We expect this technique to improve resolution and accuracy of taxonomic identification to help gain quantitative information for bulk samples of pollinators or pollens. Although there are technical challenges to be resolved, the robustness of the new methodology has been validated in relevant biodiversity studies, suggesting promise in constructing pollination networks.

Key words: mitochondria, chloroplast, pollen, metabarcoding, metagenome, PCR-free, quantify

图1

传粉网络构建流程及混合花粉组成分析方法比较。图中蓝色框和绿色框中内容分别为形态学与宏条形码方法, 橙色框中内容为宏基因组方法。底部黑色网络为真实传粉网络模型, 蓝、绿和橙色网络模型分别代表经对应方法分析可能得到的网络结构。由于各种方法都有其局限性, 所得网络均与真实网络存在一定差异。宏条形码和宏基因组技术可解决孢粉学种内形态差异的问题, 但仍存在部分近缘种难以区分的情况。相对于宏条形码技术, 宏基因组技术可避免PCR带来的物种偏倚性, 提高相对丰度的准确率。"

表1

蜜蜂科20个物种(包括蜜蜂属6个物种)列表及其线粒体基因组NCBI序列号"

物种
Species
NCBI序列号
NCBI accession number
物种
Species
NCBI序列号
NCBI accession number
Apis andreniformis KF736157.1 Bombus lapidarius KT164641.1
Apis cerana NC_014295.1 Bombus lucorum KT164681.1
Apis dorsata KC294229.1 Bombus pascuorum KT164630.1
Apis florea NC_021401.1 Bombus terrestris KT368150.1
Apis mellifera sahariensis NC_035883.1 Melipona bicolor NC_004529.1
Apis nigrocincta KY799147.1 Melipona scutellaris NC_026198.1
Bombus breviceps MF478986.1 Nomada fabriciana KT164663.1
Bombus consobrinus MF995069.1 Nomada flava KT164670.1
Bombus hypocrita sapporensis NC_011923.1 Nomada flavoguttata KT164617.1
Bombus ignitus NC_010967.1 Nomada goodeniana KT164660.1

图2

遗传距离图绘制流程"

表2

兰科84个物种(包括石斛兰属31个物种)列表及其叶绿体基因组NCBI序列号"

物种
Species
NCBI序列号
NCBI accession number
物种
Species
NCBI序列号
NCBI accession number
Anoectochilus emeiensis NC_033895.1 Dendrobium parciflorum NC_035334.1
Apostasia odorata NC_030722.1 Dendrobium parishii NC_035339.1
Bletilla ochracea NC_029483.1 Dendrobium pendulum NC_029705.1
Bletilla striata NC_028422.1 Dendrobium primulinum NC_035321.1
Calanthe triplicata NC_024544.1 Dendrobium salaccense NC_035332.1
Cattleya crispata NC_026568.1 Dendrobium spatella NC_035333.1
Cattleya liliputana NC_032083.1 Dendrobium strongylanthum NC_027691.1
Cephalanthera longifolia NC_030704.1 Dendrobium wardianum NC_035329.1
Cymbidium aloifolium NC_021429.1 Dendrobium wilsonii NC_035330.1
Cymbidium ensifolium NC_028525.1 Dendrobium xichouense NC_035341.1
Cymbidium faberi NC_027743.1 Elleanthus sodiroi NC_027266.1
Cymbidium goeringii NC_028524.1 Epipactis mairei NC_030705.1
Cymbidium kanran NC_029711.1 Epipactis veratrifolia NC_030708.1
Cymbidium lancifolium NC_029712.1 Erycina pusilla NC_018114.1
Cymbidium macrorhizon NC_029713.1 Gastrochilus fuscopunctatus NC_035830.1
Cymbidium mannii NC_021433.1 Gastrochilus japonicus NC_035833.1
Cymbidium sinense NC_021430.1 Goodyera fumata NC_026773.1
Cymbidium tortisepalum NC_021431.1 Goodyera procera NC_029363.1
Cymbidium tracyanum NC_021432.1 Goodyera schlechtendaliana NC_029364.1
Cypripedium formosanum NC_026772.1 Goodyera velutina NC_029365.1
Cypripedium macranthos NC_024421.1 Habenaria pantlingiana NC_026775.1
Dendrobium aphyllum NC_035322.1 Habenaria radiata NC_035834.1
Dendrobium brymerianum NC_035323.1 Listera fugongensis NC_030711.1
Dendrobium catenatum NC_024019.1 Ludisia discolor NC_030540.1
Dendrobium chrysanthum NC_035336.1 Masdevallia coccinea NC_026541.1
Dendrobium chrysotoxum NC_028549.1 Masdevallia picturata NC_026777.1
Dendrobium crepidatum NC_035331.1 Neottia ovate NC_030712.1
Dendrobium denneanum NC_035324.1 Neottia pinetorum NC_030710.1
Dendrobium devonianum NC_035325.1 Oberonia japonica NC_035832.1
Dendrobium ellipsophyllum NC_035340.1 Paphiopedilum armeniacum NC_026779.1
Dendrobium exile NC_035343.1 Paphiopedilum niveum NC_026776.1
Dendrobium falconeri NC_035326.1 Pelatantheria scolopendrifolia NC_035829.1
Dendrobium fanjingshanense NC_035344.1 Phalaenopsis equestris NC_017609.1
Dendrobium fimbriatum NC_035342.1 Phragmipedium longifolium NC_028149.1
Dendrobium gratiosissimum NC_035327.1 Sobralia callosa NC_028147.1
Dendrobium henryi NC_035335.1 Thrixspermum japonicum NC_035831.1
Dendrobium hercoglossum NC_035328.1 Vanilla aphylla NC_035320.1
Dendrobium huoshanense NC_028430.1 Vanilla planifolia NC_026778.1
Dendrobium jenkinsii NC_035337.1 Sobralia aff. bouchei NC_028209.1
Dendrobium lohohense NC_035338.1 Phalaenopsis hybrid NC_025593.1
Dendrobium moniliforme NC_035154.1 Phalaenopsis aphrodite formosana NC_007499.1
Dendrobium nobile NC_029456.1 Oncidium hybrid NC_014056.1

图3

线粒体基因组和叶绿体基因组种间及属间遗传距离图。(a) Apis属内6个物种线粒体基因组上13个蛋白编码基因序列P-distance图; (b) Apidae科内20个物种线粒体基因组上13个蛋白编码基因序列P-diatance图; (c) Dendrobium属内31个物种叶绿体基因组上67个蛋白编码基因序列P-distance图; (d) Orchidaceae科内84个物种叶绿体基因组上67个蛋白编码基因序列P-distance图。图a和b中, 橙色区域为COI条形码序列所在位置, 橙色虚线圆圈表示COI条形码序列P-distance中值; 图c和d中, 橙色区域为matK条形码序列所在位置, 紫色区域为rbcL条形码序列所在位置, 橙色虚线圆圈表示matK条形码序列P-distance中值, 紫色虚线圆圈表示rbcL条形码序列P-distance中值。"

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