Radish Genetics Demystified?

[Andrew Barney]

Still not sure whether this should be in the "Greens & Brassicas" sub-forum or not. I decided probably not, though a mod can move this thread if needed.

Based on this thread by Klaus http://opensourceplantbreeding.org/forum/index.php?topic=476.0

and this thread i started http://opensourceplantbreeding.org/forum/index.php?topic=555

Dan Brisebois, the breeder of these black-skinned radishes, has written a nice blog post about their breeding history:

https://goingtoseed.wordpress.com/2019/11/15/living-the-radish-breeding-dream/. I also really love the "skunk radishes"!

I've gotten bit by the radish color genetics bug. In addition to starting the mild purple radish selection project I've also planted yellow sweet radishes next to "sichuan-red-beauty-radish". I'm wondering if crossing sichuan-red-beauty that has red flesh to my sweet purple skinned radishes might produce a purple skinned purple fleshed radish. I plan to try.

The interesting thing is that radish skin color genetics has not been explored since 1938! The best paper on google scholar is one mostly in Japanese from 1938! Enough of it is also in English. https://www.jstage.jst.go.jp/article/ggs1921/14/1-2/14_1-2_39/_pdf

The basic takeaway from the study is that they did almost every skin color cross imaginable, but i think they may have missed a few that would be interesting to do in modern times.

They did:

F2 Red x White
F2 Red x White #2 (and reciprocal)
F2 Purple x White (and reciprocal)
F2 Yellow x White
F2 Black x White
F2 Red x Yellow
F2 Red x Yellow #2
F2 Green x Yellow
F2 Purple x Yellow
F2 Purple x Red
F2 Black x Red
Backcross F1 (black x red) x red
F2 Black x Purple
F2 Black x Yellow

The results of three of the crosses were VERY interesting when compared to the others. These were:
F2 Red x Yellow #2 = Purple, Red, Yellow, White in 9:3:3:1 ratio
F2 Red x White #2 = Purple, Red, White in 9:3:4 ratio
F2 Purple x Yellow = Purple, Yellow, White in 12:3:1 ratio

It is clear from these results radish skin color genetics wants to behave in a 9:3:3:1 ratio. But in the case of Red x White there is no yellow locus available so the ratio skews in the favor of white instead. In the case of Purple x Yellow there is no red locus available so the ratio skews in favor of purple instead of red.

Trying to figure out black skin color has also been very interesting and very confusing. According to the study they came to the conclusion that black and yellow might be on the same chromosome and that is why you can never seem to get black and yellow at the same time. After reviewing the data thoroughly, this is possible, in which case i think it might still be possible to get black and yellow together if you were to cross black and yellow radishes over and over and over in order to try and break that tight linkage and get a very rare recombination. That would be very cool for someone to try.

Another possibility for black skin genetics is that black and yellow are NOT on the same chromosome, but rather how the yellow gene works. It is very likely that the way the yellow color works in radishes (as it does in many other crops) is that the yellow radish gene actually breaks all expression of anthocyanins and leaves only the expression of the carotene content behind. If this is true then black and yellow are on separate chromosomes but will never be able to be together as a yellow radish could actually be hiding a black skinned gene.

The two crosses regarding black skin color that are the most interesting are:
F2 Black x Red = BP, P, BR, R, B, W with a 27:9:9:3:12:4 ratio!
F2 Black x Purple = BP, P, B, W with a 9:3:3:1 ratio
and backcross F1 (black x red) x red = BP, BR, P, R in 1:1:1:1 ratio

Instead of getting just black, purple, red, and white the results are more interesting with two new phenotypes being "Blackish-Red" and "Blackish-Purple".

This makes me wonder if a Blackish-Yellow is actually possible or not. Additionally theoretically there should be the possibility for orange radishes. And indeed there are some rare orange radishes seen online at farmers markets in NYC and Vermont or the north-east area of the united states. One photo of these actually appears to be a bicolor orange-red. If it is indeed a bicolor it might be possible it is actually a pinkish-yellow or redish-yellow instead of a unique orange locus of the yellow gene. Though either possibility is interesting and exciting. I would love to get my hands on an orange radish to find out. Could open up a whole new phenotype of radish!

I will follow-up this post soon with the relevant genotype information and any problems i may have found with the 1938 study.

[Adrian]

A orange radish is  an hybrid between yellow and red?
I think thr yellow color is a dominant color.

[Andrew Barney]

A orange radish is  an hybrid between yellow and red?
I think thr yellow color is a dominant color.

Do orange radishes actually exist? Not sure. And if they do are they a combination of yellow and red? Or pink and yellow? I do not know. Yellow is supposed to be recessive to purple in a 12:3:1 ratio and recessive to red in a 9:3:3:1 ratio. Yellow is supposed to be dominant over white in a 3:1 ratio.

Are you saying this is not true? Or there is an alternate yellow reddish gene available that is dominant?

[Adrian]

I don't know i have grow multicolor radish together and i have see the yellow appeard often in the hybrids in the next generation.

[Andrew Barney]

Getting this thread back on track...

I need to re-read the papers and write some of my thoughts down on paper, but It looks like the following radish colors are more or less possible:

(Listed in order by guessed dominance)

Black-Purple
Black
Black-Red
Purple
Red
Green
Yellow
White

Judging by the crossing data above I would guess that Red is actually produced by two separate genes, yellow & purple, on top of each other. If you have one white gene that cancels out the anthocyanins then you get yellow, if you get a white gene that cancels out the flavinoids you get purple, and if you get two white genes you get white. Black is probably on a separate chromosome than purple, which if true is a bit odd, but does explain why you can get black-red, black-purple, AND just black.

Still think black x yellow would be a worth while project if one puts in many back crosses to see if tight gene linkage can be broken and recombined.

This suggests that in order to get an orange radish you might need a pink radish crossed with a yellow one.

[Andrew Barney]

The probable genotypes of root colors in radish were assumed as follows from the Japanese 1925 study:

rrBBYiYi   Black
RRBByy   Purple
RRbbyy   Red
RRB_YY   Yellow
rrBByy   White

But these don’t quite make sense to me. I think they are probably wrong in some aspect or all aspects.

Also not sure about green skin color. Green is supposed to be dominant over yellow.

My guess is that the genotypes are closer to this:

BBPPYY   Black-Purple
BBppYY   Black
BBPPyy   Black-Red
bbPPYY   Purple
bbPPyy   Red
bbppyy   Yellow
bbppYY   White

If I am correct, then Black and Yellow are NOT on the same chromosome, but black is just so powerful that the yellow is always covered up. But Black does seem to be different than the Purple gene. Neither of these consider the differences between skin and flesh colors either, so this is just the tip of the ice burg i imagine.

Pink radishes are probably a modifier gene for purple. Pink would probably be something like: (bb) + (PP) + (pbpb) + (YY). If that is true, then Pink x Yellow would equal in the F1: (bb) + (Pp) + (PBpb) + (Yy) Purple. The F2 would be: bb + (PP or Pp or pp) + (PBPB or PBpb or pbpb) + (YY or Yy or yy). To get an orange in the F2 it would be something like 2/27? Might have done my math wrong, but either way it is very low numbers. Somewhere around 1 or 2 out of 30 plants. If i did my math wrong it might be out of 81 total plants.

[Andrew Barney]

Found this today. Brief study from 2019. Doesn't really help much though.

https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6764292/

[Klaus Brugger]

Very interesting topic!

I've tried to find some information about the inheritance of flesh color a while ago. There are some Chinese papers like Zhang (2006) or Lv et al. (2015), but I cannot find or access all of them.
Chen et al. (2016) have found higher expression levels of a few genes in intensely red radish.

One thing I think we have to consider is that modes of inheritance might not be the same in all cross combinations. Radish seems to have been domesticated at least three times with small European radishes and black radishes apparently being of different origin, having quite different cytoplasms (see Yamane et. al, 2009 and Li et al., 2021). This might be an explanation why Dan Brisebois had purple flesh in his black × watermelon radish F₁ but my purple small × watermelon radish F₁ didn’t show any anthocyanin in the flesh.

I must say I have very little personal experience (or theoretical knowledge!) with yellow radishes, so please don’t give the following thoughts too much weight … 
I’m not sure what actually causes the yellow color. It seems to be associated with a certain structure of the skin, not unlike the black color. In fact, I wonder if yellow, brown, grey and black might essentially be the same character. Take, for example, a brownish cultivar like the open-pollinated German 'Fetzers Maindreieck' (picture). To me, it looks somewhat like an intermediate between yellow and black.
The yellow color of radishes seems to be different from the yellow color of turnips, where yellow-skinned bulbs can have a quite intensely yellow flesh too. Maybe I’ll cross a black and a yellow turnip this year. Of course, radish and turnip are also somewhat crossable. And maybe introducing genes from yellow-flowering Raphanus raphinistrum into cultivated radish would be interesting, too.
The plants on the NYC farmer’s market look like beets to me (as has already been mentioned in the other thread), with a nice combination of different betalains ('Touchstone Gold' looks similar). However, I think an orange radish (a yellowish structure on top of a red or pink cortex …) is a very interesting breeding goal.
 
By the way, I just found a cool paper by Chen et al. (2021) where they show that anthocyanin accumulation may be enhanced by inducing tetraploidy. White tetraploid radishes of cultivar 'Rex' are commonly grown in home gardens, so tetraploidy definitely works for vegetable radishes.




Literature:

Chen, F. B., Xing, C. Y., Huo, S. P., Cao, C. L., Yao, Q. L. & Fang, P. (2016). Red Pigment Content and Expression of Genes Related to Anthocyanin Biosynthesis in Radishes (Raphanus sativus L.) with Different Colored Flesh. Journal of Agricultural Science, 8(8), 126-135.
https://doi.org/10.5539/jas.v8n8p126
Chen, F., Gao, J., Li, W., Liu, Y., Fang, P., & Peng, Z. (2021). Colchicine-induced tetraploidy influences morphological and cytological characteristics and enhances accumulation of anthocyanins in a red-fleshed radish (Raphanus sativus L.). Horticulture, Environment, and Biotechnology, 62(6), 937–948.
https://doi.org/10.1007/s13580-021-00363-w
Li X, Wang J, Qiu Y, Wang H, Wang P, Zhang X, Li C, Song J, Gui W, Shen D, Yang W, Cai B, Liu L, Li X. SSR-Sequencing Reveals the Inter- and Intraspecific Genetic Variation and Phylogenetic Relationships among an Extensive Collection of Radish (Raphanus) Germplasm Resources. Biology. 2021; 10(12):1250.
https://doi.org/10.3390/biology10121250
Lv, F. S., Tao, H. Y., Tan, G. X., & Zeng, X. X. (2015). The characteristics of parents and seed production techniques of red radish “Yanzhihongyihao”. Shanxi Agricultural Science, 61, 122-123.
Yamane, K., Lü, N., & Ohnishi, O. (2009). Multiple origins and high genetic diversity of cultivated radish inferred from polymorphism in chloroplast simple sequence repeats. Breeding Science, 59, 55-65.
https://www.jstage.jst.go.jp/article/jsbbs/59/1/59_1_55/_pdf/-char/ja
Zhang, L (2006). Inheritance of main botanical characters in radish (Raphanus sativus L.). China Vegetables, 10, 10-12.

[Andrew Barney]

Very interesting thoughts Klaus,

You may be right. I did not know about the three independent domestication events,  but I'm not entirely surprised. It is possible there are multiple pathways or alleles for various purple or red anthocyanins. Could be the same for yellow. Seems very few radish breeders exist outside China or Japan and even less have worked with yellow radishes.

Golden Helios from rhshumway looks much brighter yellow than the other yellow radishes I've seen online.

The original study said they thought yellow and black were on the same allele on the same chromosome,  so your comments make that sound possible. Still odd to me, but no reason why it couldn't be true.

The paper I linked to above did mention this:

four RsMYB1.1 homologs (RsMYB1.1-1.4) were found in “XYB36-2” radish. RsMYB1.1 and the previously mapped and cloned RsMYB1.4 (contributing to red skin) were located on different chromosomes and in different subclades of a phylogenetic tree; thus, they are different genes.

This suggests that there are at least four anthocyanin genes at play. Whether this is only skin colors or includes flesh colors is not apparent to me.

But one possible explanation is that Black, Purple, Red, Pink (modifier gene?) are all separate genes or alleles.

It actually makes sense to me that red and purple are probably separate genes independently inherited.