Scribd Logo
Upload

Welcome to Scribd!

  • 5 views

    BT30 - N - Lesson 10 - PPT

    Uploaded by

    kunal113333
    The document discusses anther, microspore and endosperm culture techniques. It describes the steps involved in each technique and factors affecting them. It also mentions some applications of these techniques like haploid and triploid plant production.

    Copyright:

    © All Rights Reserved
    Download as PDF, TXT or read online from Scribd

    BT30 - N - Lesson 10 - PPT

    Uploaded by

    kunal113333
    5 views17 pages
    The document discusses anther, microspore and endosperm culture techniques. It describes the steps involved in each technique and factors affecting them. It also mentions some applications of these techniques like haploid and triploid plant production.

    Original Description:

    Biotechnology

    Original Title

    BT30_N_Lesson 10_PPT

    Copyright

    © © All Rights Reserved

    Available Formats

    PDF, TXT or read online from Scribd

    Did you find this document useful?

    Is this content inappropriate?

    The document discusses anther, microspore and endosperm culture techniques. It describes the steps involved in each technique and factors affecting them. It also mentions some applications of these techniques like haploid and triploid plant production.

    Copyright:

    © All Rights Reserved
    Download as PDF, TXT or read online from Scribd
    Download as pdf or txt
    5 views17 pages

    BT30 - N - Lesson 10 - PPT

    Uploaded by

    kunal113333
    The document discusses anther, microspore and endosperm culture techniques. It describes the steps involved in each technique and factors affecting them. It also mentions some applications of these techniques like haploid and triploid plant production.

    Copyright:

    © All Rights Reserved
    Download as PDF, TXT or read online from Scribd
    Download as pdf or txt
    of 17

    DESIGNED AND DEVELOPED UNDER THE AEGIS OF

    NAHEP Component-2 Project “Investments In ICAR Leadership In Agricultural Higher Education”


    Division of Computer Applications
    ICAR-Indian Agricultural Statistics Research Institute
    Course Details

    Course Plant Tissue Culture and its Applications


    Name

    Lesson 10. Anther, Microspore and Endosperm Culture

    Disclaimer : Presentations are intended for educational purposes only and do not replace independent
    professional judgement. Statement of fact and opinions expressed are those of the presenter individually and are
    not the opinion or position of ICAR-IASRI. ICAR-IASRI does not endorse or approve, and assumes no responsibility
    for the content, accuracy or completeness of the information presented.

    1
    Created by

    Name Role University

    Bihar Agricultural University,


    Hidayatullah Mir Content Creator
    Bhagalpur

    Dr. Rajendra Prasad Central


    Bishun Deo Prasad Course Reviewer
    Agricultural University, Samastipur

    2
    Introduction

    • Anther culture: In vitro culture of immature anthers in which the microspores is confined
    within the anther sac at the appropriate stage of pollen development from a young flower bud.

    • Microspore or pollen culture: In vitro culture of male gametophytic cells, i.e., microspores
    or immature pollen.

    • Anther culture has an advantage over microspore culture in being very quick and the anther wall
    acts as a conditioning factor for the developing microspores.

    • But the microspore culture is preferred over anther culture as in anther culture diploid plants may
    be observed due to the involvement of tissue present in the anther.
    Anther and
    microspore culture

    Androgensis for haploid production


    Source: https://www.toppr.com/ask/question/haploid-plant-cultures-can-be-obtained-from/
    Anther culture

    Source: https://biologyreader.com/anther-culture.html
    microspore culture

    Pollen culture

    Source: https://www.biologydiscussion.com/plant-tissues/p...
    Endosperm
    culture

    Immature endosperm culture


    Mature endosperm culture
    It is the in vitro development of
    It is the in vitro development of isolated immature endosperm
    isolated mature endosperm from isola t e d a t p r e c i s e s t a g e f r o m
    ripen endospermic seed on suitable immature seed, cultured on suitable
    culture medium to obtain triploid culture medium to obtain triploid
    plantlet is called mature endosperm plantlet is called immature
    culture. endosperm culture.
    Endosperm culture

    Endosperm

    Source:http://eagri.org/eagri50/GPBR311/lec12.pdf
    Steps of anther
    culture

    1. Growing of donor plants.


    2. Selection of appropriate flower buds.
    3. Surface sterilization of the flower buds using a disinfectants (70% ethanol, tween 80 and HgCl2).
    4. Anthers along with their filaments are excised under aseptic conditions.
    5. The anthers are crushed in acetocarmine, to test the stage of pollen development stage.
    6. The anthers are detached from filaments and are placed horizontally in the culture medium and is
    then maintained with alternating periods of light (12-18 h; 5000-10000 lx m2) at 28℃ and darkness
    (12-6 h) at 22℃.
    7. As the anthers respond, wall tissues turn brown and after 3-8 weeks, they burst open due to the
    pressure exerted by the growing pollen callus.
    8. Then the embryos germinate, and are transferred to another media for development of shoot and root.
    9. For acclimatization, the plantlets are transferred to green house.
    Steps of microspore
    culture

    1. The anthers are collected from sterilized flower buds in basal media (MS or White or Nitsch).
    2. The microspores are then squeezed out of the anthers.
    3. The homogenized anthers are then filtered through a nylon sieve and the anther tissue debris is
    removed.
    4. The pollen suspension is then centrifuged at low speed (500-800 rpm/min) for 5 minutes, the
    supernatant is discarded and pellet of pollen is suspended in fresh liquid medium.
    5. Then pollen suspension is pipetted out and is spread in on a petri-dish.
    6. Petri-dishes are then incubated at 27-30°C under low intensity of white cool light (500 lux, 6 hr).
    7. Young embryoids can be observed after 30 days.
    8. The embryoids ultimately give rise to plantlets, which is later acclimatized in green house.
    Steps of endosperm
    culture

    1. Selection of explant source (mature or immature endosperm) at proper stage.


    2. The seeds are decoated, surface sterilized and then placed over a nutrient medium.
    3. The seeds are then dissected under aseptic conditions and endosperms along with embryos are
    excised.
    4. Excised endosperms are then cultured on a suitable callus induction nutrient medium and
    embryos are removed after initial stage.
    5. To induce callus, the endosperm cultures are maintained darkness or diffused light.
    Differentiation take place when calli are transferred to bright light (2000 lux – 4000 lux) and
    25⁰C temperature.
    6. Shoot bud differentiation or embryogenesis: Plantlet formation from endosperm callus
    maturation follow organogenic or embryogenic mode of development.
    7. Plantlets developed are then hardened in green house.
    Factors influencing anther and
    microspore culture
    1. Genotype of donor plants: The genotype of the donor plant plays a significant role in determining the frequency
    of pollen production and response to the culture of anther and pollen.
    2. Age of plants: Flowers from relatively young plants at the beginning of flowering season are most responsive to
    anther or microspore culture.
    3. Stage of the microspore: Anthers are more productive when cultured at late uninucleate microspore stage.
    4. Culture medium: Culture medium with activated charcoal has positive effect on the induction of androgenesis.
    The iron in the medium also stimulates the induction of haploids.
    5. Physical factors: Physical factors like light intensity, photoperiod, temperature, etc. The low temperature treatment
    to anther or flower bud enhance the haploid formation.
    6. Physiological status of donor plant: Physiological status of donor plant such as plants starved of nitrogen and
    water may give more responsive anthers compared to those that are well fed with nitrogenous fertilizers and water.
    7. Cold, hot and chemical pre-treatment
    Factors influencing
    endosperm culture

    1. Explant stage: Proper stage may vary from cellular (immature) to mature endosperm depending
    upon species. (4-7 DAP in rice, 8 DAP in wheat and 12 DAP in maize and fully matured in
    Santalum album).
    2. Nutrient medium: Undefined source like tomato juice, yeast extract, etc. can be used as nutrient
    media. Low amount of reduced nitrogen is required for proliferation of the culture.
    3. Physical factors: The pH 7.0 seems to be effective for fresh weight increase. Maximum growth
    of endosperm occurs between 24- 27⁰C temperature and 12-16 hours photoperiod with diffuse
    day light supported callusing as well as regeneration.
    4. Embryo factors: Association of embryo tissue in the initial stages seems essential for inducing
    proliferation of mature endosperm tissue in some families. Immature seeds of non-endospermic
    seeds exhibit no dependence on embryo factor.
    Applications of anther and
    microspore culture

    1. For production of haploids and homozygous plants after diploidization.

    2. For plant breeding and crop improvement: Production of haploids and homozygous diploids can be
    achieved in a smaller time period, which is time consuming through the conventional breeding
    techniques.

    3. For mutation study, microspore are advantageous for transformation because of single cell.

    4. Haploids are used in molecular biology and genetic engineering. Microspore and anther derived
    embryo and doubled haploids can be used as a starting material for transformation.

    5. To obtain the secondary metabolites. Eg. Hyoscyamus niger obtain by anther culture having higher
    alkaloid content.
    Applications of
    endosperm culture

    1. Triploid plants: Techniques of endosperm culture has enabled the production of triploid plants.
    Triploid plants are self-sterile and usually seedless.

    ü Triploidy increases edibility of fruits and is desirable in plants such as apple, banana, grape,
    watermelon, cucumber and mulberry which are commercially important.

    ü In timber and fuel yielding plants also triploids show better performance over their relative
    diploids or tetraploids.

    2. Endosperm as nurse tissue: Endosperm can be used as nurse tissue for raising hybrid embryos.

    You might also like