A RECENTLY DISCOVERED ALBINO SEEDLING MUTANT IN MAIZE

– A lethal albino seedling mutant of maize that is resistant to imazethapyr was found in the material from the Maize Research Institute, Zemun Polje, gene bank, The mutation was characterized by a normal development of the first two leaves, with white coloration appearing from the third to the fifth leaves, at the base of the leaves spreading towards their end, resulting in the death of the whole plant. While the upper leaves were dying, the first two were still green for eight to ten days. It seemed that the mutation was conditioned by one recessive allele.


INTRODUCTION
Albino mutants are well known in plant and animal species.They are usually caused by one recessive allele, which in a homozygous state leads to a total or partial absence of pigment in the organism.In maize, these are so-called white genes, and these are abbreviated by the letter w.The following alleles exist: w1, characterized by yellow kernels that germinate normally to produce lethal off-white seedlings; w2 -allelic to dek21, white seedlings, endosperm pitted and mosaic for colored and colorless aleurone when colored aleurone genes are present, plastid DNA reduced; w3 -kernels are viviparous, embryo, scutellum and endosperm are white, aleurone with A1, C1, R1, etc., is colored but with reduced pigment intensity, seedlings are white, a pale yellow kernel appearance in some backgrounds seems to be due to an unidentified flavonoid pigment; allele w3-8686 is dormant with white endosperm, pale green seedlings in dim light, blocked in the carotenoid biosynthetic pathway; w11, w14, w16 and w17 -lethal seedlings like w1; w15, like w1, which fail to convert protochlorophyllide to chlorophyllide; w18 -white seedlings, pale green streaks in some backgrounds; and w19 -white plant tissue in leaf and sheath chimeras (Neuffer et al., 1997).A similar group of genes, socalled white luteus (abbreviated as wlu: wlu1, wlu3, wlu4 and wlu5) have pale yellow lethal seedlings, varying from nearly white to yellow depending on genetic background; wlu2 has white seedlings with faint yellow green on the midrib and leaf sheath.A series of similar mutations, but some of them with mostly yellow seedlings, are designated as cl, l, lw, grt, oy and ppg.The mutant we found was not like any of those given above.Here we only describe it, but for a definite genetic description a test for allelism with similar ones would be needed.

MATERIALS AND METHODS
In 2001, eleven total and broad-spectrum herbicides were applied to the whole material of the Gene Bank of the MRI Zemun Polje-Belgrade, in order to search for potential rare mutants resistant to some of them.Only the resistance to the herbicide imazethapyr (Pivot) was unequivocally observed (Vančetović et al., 2004).Six of about 30 plants that showed the full resistance to Pivot were out-crossed to the non-resistant testers.In the year of 2002, testcrosses of the six resistant plants to nonresistant testers were selfed in order to investigate the mode of inheritance of this trait.In 2006 (the  material was planted in 2003, but a strong storm destroyed everything), F 2 generations obtained in this way were sown with as many plants as possible.They were sown ear-to-row, and unusual albino plants were observed in the progenies of some of the selfed ears of two of the resistant plants (Vančetović et al., 2007).

RESULTS AND DISCUSSION
The first two leaves in the albino plants developed normally, but a white coloration appeared from the third to fifth leaf, starting at the bottom of the leaf and spreading towards the edges, causing the death of the white leaves.The first two leaves lived for another eight to ten days.We called this phenomenon "delayed albinism" (Figs. 1 to 8).The field trial was treated only with Banvel-DP (dicamba + dichlorprop) herbicide for broad-leafed weeds which did not affect maize, and so the phenomenon was considered to be of a purely genetic nature.There is, however, a possibility that Banvel-DP (dicamba + dichlorprop) was a sort of a trigger for the expression of this genetically determined trait.However, the proof against this statement is that the plants were in different stages during the treatment with Banvel-DP (dicamba + dichlorprop), as sowing had been done by hand and the time of plant emergence varied due to a very cold spring (plants were in the 3-5-leaf stage), but all the plants showed the same symptoms (occurring at the same spot on the plant).
In order to investigate the mode of inheritance of the trait, the albino and normal green plants in the F2 generations were counted, and the χ 2 test for the segregation of a 3:1 ratio of green:albino plants was performed.The obtained results were perplexing (Table 1).
Since not all progenies of the selfed ears in the F2 generations of plants no. 2 and 5 (resistant to imazethapyr (Pivot)), showed segregation for albino plants (four not segregating ears versus five segregating ears for plant no.2, and two not segregating ears versus three segregating ears for plant no.5), and since the involved sister cross-testers and their components never showed albinism, it was presumed that plants 2 and 5 were heterozygous for the respective albino gene(s) which expressed their phenotype in the examined F2 generations.As the phenomenon was the same for the progenies of plants 2 and 5, they had to be genetically closely related, which was also supported by their mutual resistance to the herbicide (Vančetović et al., 2007).These plants probably originated from the same line or population (mixtures of our gene bank accessions comprised from 20 seeds of each population and 10 seeds of each inbred line, for the test in 2001 (Vancetović et al., 2004)).It was also assumed that the seeds of the albino plants could have a lower percent and/or emergence energy in comparison with the seeds of the normal green plants.This could be the reason for a lower number of albino plants in the progenies of plant no. 2 than theoretically expected.For this reason the emergence for each segregating F2 population was calculated, but no obvious corre-lation between emergence and the obtained segregation ratio (Table 1) was found.
Furthermore, the χ 2 test was separately performed for plants 2 and 5, summing over all cobs and F2s.Results are given in Table 2.The expected ratio of 3:1 of green : albino plants was registered in plant no. 5, while this was not the case with plant  no. 2. Given that the total number of plants and the emergence percentages were much higher for plant no. 5 than for no.2, the obtained results for plant no. 5 are probably more statistically accurate than for plant no. 2.
In 2006 as many green plants as possible were selfed in the segregating populations and in 2007 the expression of this trait was observed.The selfed material was sown ear-to-row with 36 plants in each row for all selfed F2s.It was sowed on an area without herbicide treatment, so that a conclusion on the effects of Banvel herbicide on this phenomenon could be drawn.
As in the previous year the albino plants appeared in some of the selfed progenies, so the influence of Banvel was excluded.Green versus albino plants in the progenies, as well as segregating versus all-green progenies in all selfed F2s, were counted.Results of χ 2 tests for the 3:1 segregation ratio in the selfed progenies per se, and also for their sum in a particular F2 (because of a small number of plants in the selfed progeny per se) are given in Table 3. Table 4 shows the χ 2 values for the 3:1 segregation ratio separately for a plant number of both 2 and 5, for all selfed F2 generations, while Table 5 presents χ 2 values for the segregation ratio of 2:1 of segregating versus all-green selfed progenies in a particular F2.None of the χ 2 values was significant.Based on these results it is clear that this trait is conditioned by one recessive allele of an actually dominant gene.

Table 2 .
χ 2 test for the segregation of 3:1 ratio of green : albino plants in segregating F2 populations separately for plant 2 and 5, respectively.

Table 1 .
χ 2 test for the segregation of 3:1 of green : albino plants in segregating F2 populations.
a Genotypes are obtained by out-crossing plant no. 2 and plant no. 5 resistant to Pivot to the non-resistant sister-cross testers (T), and /1, 2…is a designation of the selfed plant.b χ 2 values for 1 DF: probability 0.05=3.841;0.01=6.635.

Table 3 .
Results of χ 2 tests for the 3:1 segregation ratio in selfed progenies per se, and also for the sum of them in a particular F2