Chorion-deficient Mutants ?

Graham Corley-Smith corley at
Sun Oct 23 17:09:20 EST 1994

RE: message of 23 Oct 94 15:44:18 -0230 from pvalerio at
   regarding possibility of making chorion-deficient mutants.

Hi all,
   I find the concept of producing chorion deficient mutants intriguing.
However, I was under the impression that the
chorion is extremely important in fertilization for reasons most of you I
am sure are familiar with.  For example, if the chorion was lost, would not
also the micropyle be lost?  Therefore, losing a primary block to
polyspermy?  Is anyone aware of experiments where the chorion has been
removed before fertilization and viable progeny have been produced?
P.S.  In hope that it may help save time to anyone that decides to
      investigate chorion deficient mutants or mutants that have softer
      chorions, I have attached below, a list of a few references that
      may help.

Paul.Becker, K. A. and N. H. Hart (1989) Polar body formation in
  zebrafish eggs.  Annual meeting of the American Society
  of Zoologists, American Microscopical Society, Animal
  Behavior Society, Crustacean Society, International
  Assoc. of Astacology, and the Society of Systematic
  Zoology, Boston, MA, USA, December 27-30, 1989.  Am.
  Zool. 29, 108A. [4]
Becker, K. A. and N. H. Hart (1990) Stage-specific effects
  of cytochalasin D on polar body formation in the teleost
  egg.  13th annual meeting of the American Soc. for Cell
  Biology, San Diego, CA, USA, December 9-13, 1990.  J.
  Cell Biol. 111, 111A. [5]
Donovan, M. J., Hart N. H. (1986). Cortical granule
  exocytosis is coupled with membrane retrieval in the egg
  of Brachydanio.  J. Exp. Zool. 237:391- 406. [13]
Donovan, M. J., Hart, N. H. (1983). Sensitivity of the
  cortical reaction to cytochalasin B in eggs of
  Brachydanio-rerio.  Am. Zool. 23:984. [14]
Hart, N. H. (1970). Artifical activation of the ripe egg in
  Brachydanio.  Anat. Rec. 166:315. [19]
Hart, N. H. (1987). Morphological features of the plasma
  membrane during activation of the Brachydanio egg.  J.
  Cell. Biol. 105:254A. [20]
Hart, N. H., Collins, G. C., Wolenski, J. S. (1986).
  Ultrastructure of the egg cortex of Brachydanio.  Am.
  Zool. 26:11A. [21]
Hart, N. H., Donovan, M. (1983). Fine structure of the
  chorion and site of sperm entry in the egg of Brachydanio
  rerio.  J. Exp. Zool. 227:277-296. [22]
Hart, N. H., Messina, M. (1972). Artificial insemination of
  ripe eggs in the zebrafish, Brachydanio rerio.  Copeia
  2:302-305.  [131]
Hart, N. H., Wolenski, J. S., (1989). Actin in the cortex of
  the teleost egg.  J. Cell Biol. 107:174a. [23]
Hart, N.H. and G.C. Collins (1991) An electron-microscope
  and freeze-fracture study of the egg cortex of
  Brachydanio rerio.  Cell Tissue Res. 265, 317-328. [24]
Hart, N.H., Becker, K.A., Wolenski, J.S. (1992). The sperm
  entry site during fertilization of the zebrafish egg:
  localization of actin.  Mol. Reprod. Dev. 32:217-228.
Nugent, J., Hart, N. H., Schalkoff, M. (1981). The cortical
  cytoplasm and sites of sequestered calcium in the teleost
  egg.  Am. Zool. 21:984. [125]
Wolenski, J. S., Hart, N. H. (1987). Scanning electron
  microscope studies of sperm incorporation into the
  zebrafish (Brachydanio) egg. J. Exp. Zool. 243:259-274.
Wolenski, J. S., Hart, N. H. (1987). Sperm incorporation and
  ferilization cone formation in eggs of the zebrafish
  Brachydanio.  J. Cell Biol.  105:338A. [65]
Wolenski, J. S., Hart, N. H. (1988). Effects of
  cytochalasins B and D on the fertilization of zebrafish
  Brachydanio eggs.  J. Exp. Zool. 246:202-215. [66]
Wolenski, J. S., Hart, N. H. (1988). Sperm incorporation
  independent of fertilization cone formation in the danio
  egg.  Dev. Growth Differ. 30:619-628. [130]
Wolenski, J. S., Hart, N. H. (1988). The fertilization cone
  develops in the presence of actin filament cappers.  J.
  Cell Biol. 107:452a. [67]
Wolenski, J., Hart, N. H. (1984). Surface events at
  fertilization in the teleost egg.  Am. Zool. 4:23A. [68]
Wolenski, J., Hart, N. H. (1985). Sperm incorporation into
  the teleost egg.  Am Zool. 25:96A. [69]
Harvey, B., Kelley, R. N., Ashwood-Smith, M. J. (1983).
  Permeability ofintact and dechorionated zebra fish
  (Brachydanio rerio) embryos to glycerol and dimethyl
  sulfoxide.  Cryobiology 20:432-439. [27]
Hisaoka, K. K. (1958). Microscopic studies of the teleost
  chorion.  Trans. Am. Micro. Soc. 77:240-243. [114]

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In Message 23 Oct 94 15:44:18 -0230, pvalerio at writes:

>   I am curious to know whether anyone is currently working on, or has
>considered the possibility of exploring, the following research topic with
>fish embryos.
>   Many people working with fish embryos need to routinely remove the chorion
>as part of their experimental protocol. This can be time-consuming, and perhaps
>harmful to the embryo. Is there a possibility of creating a chorion-deficient
>mutant of, say a zebrafish, analogous to the cell-wall deficient algal cells
>that are used by plant molecular biologists ? The rationale behind this
>procedure is as follows:
>1. Mechanical or enzymatic dechorionation can be harmful to
>    embryos.
>2. Although the chorion may be required for osmoregulation
>   (eg. by retaining colloid released during cortical
>    granule breakdown), dechorionated embryos can have similar survival
>    rates in freshwater (+ marine ?) fishes.
>3. Chorion-deficient embryos could be used for a variety of experiments
>   that are difficult or impossible with normal embryos: eg. cell fusion,
>   gene transfer, organelle or cell ablation by lasers, other forms of
>   microsurgery, etc...
>4. Perhaps chorion-deficient embryos could be cryopreserved, since the
>   problem of supercooling would be circumvented by exposing the embryo
>   directly to cooled cryoprotectant solutions.
>   I understand that the chorion protein may be derived from more than one
>cell type in the ovary, but even if a mutation could be produced that prevents
>the 'hardening' process at the time of cortical granule breakdown, the
>chorions could be more easily removed. If this could be achieved with a model
>species like the zebrafish, it might make life a lot easier for people working
>with other species having a very tough chorion - if the method could be
>adapted to work with other species. At the moment, many studies must involve
>more time-consuming procedures, such as microinjection through the micropyle,
>for example.
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                        Graham E. Corley-Smith
                        Institute of Molecular Biology and Biochemistry
                        Biological Sciences Department
                        Simon Fraser University
                        Burnaby, B.C.,
                        V5A 1S6
                        Phone: (604) 291-3021
                        Fax:   (604) 291-5583
                        e-mail: corley at

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