Chickens You Can’t Count On1
Page 1 - Hens rescued from the pot
In the early 1950’s Mr. F. H. Leuscher, an extension poultry specialist in
Pennsylvania, reported that a commercial poultry breeder near Williamsport was
experiencing unusual embryonic mortality in some pedigree matings of his
Single-Comb White Leghorn stock. Regardless of the sire, none of the eggs
from certain hens hatched. Mr. Leuscher also noted that the egg white from
these eggs seemed to lack a yellowish color typical of most eggs. He brought
three of these hens to Prof. A. J. C. Maw in the Poultry Science Department at
Penn State for further study.
If you were Prof. Maw, what could you conclude from the information provided?
What questions would you have?
How would you figure out the problem with the hens/eggs?
Single Comb White Leghorn
http://www.agric.gov.ab.ca/images/livestock/aprc/rarebreed_scwl.jpg
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PBL Problem written by H. B. White, Department of Chemistry and Biochemistry, University of Delaware, Newark, DE.
19716, (halwhite@udel.edu) November 2002, revised 19 December 2002.
Chickens You Can’t Count On1
Page 2 - What color is egg white?
Professor Maw knew that the yellowish color of normal chicken egg white was
due to Vitamin B2, also known as riboflavin. He suspected that the eggs were
riboflavin deficient so he fed the his three hens a diet supplemented with a tenfold excess of riboflavin, mated them with normal roosters, collected the eggs,
and incubated them. None of the eggs had a yellowish egg white and none of
the eggs hatched. Then he decided to inject riboflavin directly into the egg white
before incubation. He punched a small hole above the air sack in the blunt end
of a dozen eggs, injected a sterile solution of FMN (a soluble riboflavin
derivative), covered the hole with Duco cement, and incubated the eggs.
Twenty-one days later he had twelve downy chicks, seven pullets and five males.
In his next experiment, Dr. Maw mated the siblings and incubated the eggs
without injecting FMN.
Predict what happened.
1
PBL Problem written by H. B. White, Department of Chemistry and Biochemistry, University of Delaware, Newark, DE.
19716, (halwhite@udel.edu) November 2002, revised 19 December 2002.
Chickens You Can’t Count On1
Page 3 - Fluorescent “egg”citement
To Prof. Maw’s surprise, all of the eggs from the F1 dihybrid cross hatched. Among the F2
females that hatched, 21 out of 88 subsequently laid eggs that lacked a yellow-colored egg white
and would only hatch if supplemented with riboflavin. Prof. Maw concluded that an autosomal
recessive gene (rd for riboflavin deficient) was responsible for the inability to deposit riboflavin.
The mutation attracted the attention of other Penn State researchers−Edward Buss, a poultry
geneticist, and Carl Clagett, a biochemist, who were interested in how normal hens are able
A riboflavin solution
fluorescing under an
ultraviolet light.
to put riboflavin in their eggs and what was wrong with the mutant hens.
After several frustrating years of testing and rejecting various hypotheses
associated with abnormal excretion of riboflavin by the kidneys (avian
riboflavinuria) in the mutant strain, serendipity struck. Bill Winter, a
graduate student, was trying to develop a simple assay for riboflavin in egg
white from normal and mutant eggs. He was puzzled by the fact that
riboflavin, which normally is fluorescent under ultraviolet light (picture at
left), did not fluoresce in normal egg white. In fact, when he added
riboflavin to diluted normal egg white, the fluorescence was quenched.
However, after several additions, the fluorescence no longer was
quenched. In contrast, when he added riboflavin to the diluted mutant egg
white, there was no quenching.
A representation of the experiment is shown below.
Fluorescence
Mutant egg white, rdrd
Normal egg white, RdRd
Volume of riboflavin solution
What are the implications of this experiment?
1
PBL Problem written by H. B. White, Department of Chemistry and Biochemistry, University of Delaware, Newark, DE.
19716, (halwhite@udel.edu) November 2002, revised 19 December 2002.
Chickens You Can’t Count On1
Page 4 - Burning chicken fat
The Rd gene codes for a riboflavin-binding protein whose normal function is to
transport riboflavin from the hen to her eggs. Homozygous rdrd hens cannot
produce the protein and thus cannot deposit riboflavin in their eggs. With only
trace amounts of riboflavin, embryos in eggs laid by rdrd hens die on or about 13
days of incubation. This is a time when the chicken embryo depends almost
exclusively on the oxidation of yolk triglycerides for energy as shown by a
respiratory quotient (RQ) of 0.72.
How might riboflavin deficiency lead to embryonic death?
Energy production and respiratory quotient (R.Q.) of chicken eggs incubated at different temperatures for the first 10 days
and at 37.5 C thereafter. (From Decuypere et al 1979 as modified by Rahn and Paganelli, 1990).
1
PBL Problem written by H. B. White, Department of Chemistry and Biochemistry, University of Delaware, Newark, DE.
19716, (halwhite@udel.edu) November 2002, revised 19 December 2002.
Chickens You Can’t Count On1
Problem Resources
Buss, E. G. 1969 Genetic interference in the egg transfer, utilization and requirement of
riboflavin by the avian embryo. In The Fertility and Hatchability of the Hens Egg, Ed. T.
C. Carter, B.M. Freeman, pp. 109-16. Oliver & Boyd, Edinburgh, UK.
Chatellier, D. S. and White, H. B. III 1988 What color is egg white? A biochemical
demonstration of the formation of a vitamin-protein complex using fluorescence
quenching. J. Chem. Educ. 65, 814-5.
Maw, A. J. G. 1954 Inherited Riboflavin deficiency in chicken eggs. Poultry Sci. 33, 216217.
Rahn, H. and Paganelli, C. V. 1990 Energy budget and gas exchange of avian eggs. In
Avian Incubation, Tullett S. G. ed. pp. 175-193. Butterworth-Heinemann, London.
Romanoff, A. L. and Bauerfeind, J. C. 1942 Influence of riboflavin-deficiency in eggs on
embryonic development (Gallus domesticus). Anat. Recd. 82, 11-23.
Winter, W. P., Buss, E. G., Clagett, C. O., and Boucher, R. V. 1967 The nature of the
biochemical lesion in avian renal riboflavinuria-II. The inherited change of riboflavin
binding protein from blood and eggs. Comp. Biochem. Physiol. 22, 897-906.
White, H. B. III and Merrill, A. H. Jr. 1988 Riboflavin-Binding Proteins. Ann. Rev.
Biochem. 8, 279-99.
White, H. B. III 1991 Maternal diet, maternal proteins, and egg quality. In Egg
Incubation: Its Effects on Embryonic Development in Bird and Reptiles. Ed. M. W. J.
Ferguson and D. C. Deeming pp. 1-15 Cambridge University Press
White, H. B. III 1996 Sudden death of chicken embryos with hereditary riboflavin
deficiency. J. Nutr. 126, 1303S-1307S.
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PBL Problem written by H. B. White, Department of Chemistry and Biochemistry, University of Delaware, Newark, DE.
19716, (halwhite@udel.edu) November 2002, revised 19 December 2002.