Without further information on the phenotypes I can only think of a conditional phenotype, such as reported for the zwille (zll) mutant. In such a case, even the homozygous mutant will show several classes, which upon selfing will also segregate for the different classes just as you observed. The frequency of the different classes might change by different growth conditions.

Thanks a lot for your valuable opinion.

Following is the brief description of classes' phenotype.

The classes are categorized on the basis of plant height and flowering time on maturity. But on early stages based on rosette area or biomass and flowering time. these phenotypes are not affected by growth conditions.  Pictures are attached herewith. Kindly have a look and comment further. Looking for your guidance and experts opinion. 

regards

If you blasted your Arabidopsis with EMS, you can expect that a great number of mutations will have occurred (somewhat) randomly throughout the genome, depending on the dosage. I'm not surprised that you are observing segregation for a range of traits among the progeny.

You have defined 4 phenotypic classes in each generation. Are you certain of your classification? What were the ratios of phenotypic classes in each generation?

If your C1 class is a result of heterozygosity at one or more loci, then your selection for that phenotypic class will always result in segregating selfed progeny.

flowering can be a function of daylength and can be conditional upon the plant being physiologically mature to be receptive to the stimulas of change in day length. Floral initiation and floral induction are often influenced by environment triggers ie. daylength, vernalization. Are you sure the phenotypes are not conditioned on internal (plant) or external (environment) factors? Your phenotype classes are based on characteristics related to the physiology of flowering.  These are very sensitive mechanisms that are controlled by whole sets of genes and regulatory DNA components. I agree with Jack Erron Haggard, you should expect unusual segregation. Try to understand the ratio of phenotypes using a framework where different dominant or recessive alleles at one or more loci controlling the phenotype, perhaps with an environmental trigger. But in an undirected mutagenesis project quantitative genetics easily breaks down.

Many thanks to Jack Erron Haggard  and Cathy Wilson.

I am sure that these phenotype are not conditional. I sow and always placed all the plants in the same chamber and observed the consistent phenotypes.  But their phenotypic ration is not as Mendelian genetics reveals. How can I elaborate this case using the framework where one or more heterozygous locus could be the cause of this phenotype. Kindly suggest any book or article of similar instance.

regards

What do you mean that their phenotypic ratio is not Mendelian? You observed four phenotypic classes. How many individual progeny did you observe in each generation? How many of these fit into each class?

Did you use Chi-square or other goodness-of-fit tests? If so, what expected ratios did you test?

Have you tried isolating each phenotype (maturity, height, etc.) and breaking the progeny down into two groups (or however many groups you can distinguish...) for each and then testing their goodness-of-fit to Mendelian ratios?

I generally agree with what Cathy has said, except that quantitative genetics does not "break down" (nor does Mendelian genetics). Your first generation will be a big mess, and you may have a load of recessive lethals to clear out, which may make it seem as if genetics "broke down", but I assure you that the principles remain in place.

Total number of segregates in each category (not ratio)

C1 segregates into C1:C2:C3:C4 as 43:41:6:8

C2 segregates in to C1:C2:C3:C4 as 1:20:2:10

C3 segregates in to C1:C2:C3:C4 as 1:0:42:0

C4 segregates in to C1:C2:C3:C4 as 1:0:0:2

regards

Hi Muhammad,

1. Have you grown wild-type of the same ecotype in the greenhouse where you grew these plants as a control? Did the wild-type plants look normal? Disease can sometimes stunt growth or disfigure phenotypes of plants.

2. The C3 segregation data, all 42 offspring were classified as C3, so it should be homozygous for the trait. The unique '1' plant, probably from seed contamination when you harvested them.

3. You had only 3 seeds for C4 segregation test? Or, most of them did not germinate?

I agree with Yuan-Yeu Yau - based on these results, C3 appears to be homozygous at all loci controlling your phenotype.

I assume that C4 is the weak little plant in your images and only produced 3 seed. Such a small sample is insufficient for segregation testing (I'd prefer to see >100, but I understand the limitations of Arabidopsis).

I wouldn't compare with wild type as a control, but instead compare with the non-EMS mutagenized transformant. Does it appear to fit one of your four categories?

Yes, sorry, should compare to the non-EMS mutagenized transformants, not the wild-type.

Dear all,

Yes, every time I sow the Non-mutagenized (wild type) as control. 

The C4 type plants produced very few seeds, I bulked the 10 plants which produced almost 10 seeds, out of which only three germinated.

I crossed the C3 with non-mutagenized mother, all the F1s are normal  like non-mutagenized indicating recessive nature of mutation. But F2 generation segregates in to all four categories again. this time I did not count segregation ratio. 

This is all what I have known so far. Kindly shed light on this aspect too.

regards

This is normal. With strong mutagens multiple mutations may happen in the same cell. The derived mutant, when backcossed, yields a F2 segregating for all the mutations. I described such a case earlier, see attached publication.

Regards, Erik

@ Muhammad,

1. I am very curious, why did you use a 'transgenic' background line for EMS mutagenesis study at first place. Are these two evens (transgenesis + EMS mutagenesis) needed in your research? I am wondering how would you explain to the paper reviewers that the transgenesis background of your materials have NOTHING at all to do with the current 4 phenotypes if they ask.

2. Do you intend to find out the mutation site(s) by EMS?

Dear Yuan-Yeu Yau ·

1) I used the GUS as marker driven by promoter of specific gene and this mutant was not seen have any direct role for that gene. But when I sown the M3 generation along with non-mutagenized progenitor, this mutant segregates into different morphological classes of plants. The non-mutagenized progenitor have no difference as compared to its genetic back-ground non-transgenic Col-0 wild type. So, these phenotypes are purely the result of EMS induced mutations.

2) At this moment I just want to know the possible genetic causes for the inheritance of these phenotypic classes.  

regards

you have to assessment the morphological traits in vivo under stress.