PLS221                                        Instructor: Carlos F. Quiros

Amarillidaceae (Liliaceae): Allium cepa and allies

List of references (includes reading assignments) 

Slides (onion, garlic, asparagus)


Onions and their allies:

Genus Allium, genomic numbers

New World and Old World Alliums

Main Allium crops, their origin distribution and domestication

Onion: A. cepa

Horticultural types, bunching onions

Interspecific hybridization

Cytogenetics, addition lines

Floral Biology

Male sterility

Inheritance of most important traits:

Bulb color

Diseases

Mapping and molecular markers

Biotechnology

Other cultivated species



Genus Allium encompass about 600 species.

Includes onions, garlic, leek, and others.

Bulbous biennial or perennial herbs, which give off distinctive and pungent odor when tissues are crushed. Onion smell is the most distinctive diagnostic feature of the genus.

Genomic numbers range from x=7, 8, 9, and 10

Alliums are widely distributed through out the temperate Northern Hemisphere. In the Old World, Allium species  abound in Europe, particularly USSR, N. Africa and Asia. The range of the genus embraces the Old World centers of early agriculture in Western Asia and China, but it does not reach down as far as ancient American centers of civilization. In the New World there are approximately 80 different species, mostly in Western USA, extending to Mexico, and Guatemala.

A. schoenoprasum (chives) is common to both N. America and Eurasia, from the Arctic to tropical latitudes!

With the exception of some species of chives, all the cultivated species have x=8. The genus includes species that outcross, and species that are sterile and reproduce asexually.
 

Genome evolution:

Old world species are mostly x=8, only 10% are x=7

90% of the New world Alliums are x=7

The majority of the species are diploids with x=8, followed by x=7 species, and a few with x=9 chromosomes.

x=10 is considered as a deviant number, restricted to a few Russian species with 2n=2x=20.

Polyploidy is common in the family, ranging from diploids to octoploids. Most often these are autopolyploids, although some 2n=4x=48 and 2n=4x=32 allopolyploids have been reported.

It is unknown which is the ancestral genomic number, although it is assumed to be x=8 because this is the most common number, thus x=7 and x=9 might derive from it.

Other evidence points to x=7 as the basic genomic number. This evidence centers on karyotype symmetry. x=7 chromosomes are symmetric with median centromeres, while those in x=8 and 9 have submedian centromeres. There is documented evidence of centromere misdivision generating asymmetric genomes by Robertsonian translocations. In any case, evolution of the Allium genomes has involved chromosomal rearrangements, such as inversions and translocations, and aneuploidy.

Other evidence based on phytogeography indicates that North American species with x=7 migrated to this continent before the Cenozoic era when Alaska and Siberia were bridged by land, thus surviving the Ice Age. Most of the x=7 species left in Europe were extinguished (only 10% of Alliums in Europe have x=7). The predominant x=8 species re colonized Eurasia.

B chromosomes: These are accessory or supernumerary chromosomes, which do not have homology to the basic genome. They have small size, are unstable, display numerical polymorphism and do not seem to affect the plant phenotype. They appear in 5% of Allium species. Their origin remains a mystery. They are mitotically unstable, but do not have a known mechanism of accumulation as in maize. There is polymorphism within B chromosomes, some are telo, other metacentric, at least 29 different karyotypes. They undergo regular pairing and chiasmata at meiosis.

Karyotypes and DNA content: Because of differences in karyotype morphology, it is evident that karyotype evolution in Allium has involved a minimum of structural rearrangements specially detectable in nucleolar organizing chromosomes. Usually two pairs of chromosomes in the genome organize nucleoli, displaying satellites. Changes in structure of this chromosome have been used to study the evolution in the genus. Changes are believed to be due to inversions and chromatin exchanges. Different types have been defined depending on the species where each type is found.

C-banding is also quite useful in Allium for this type of studies.

Chromosome size varies widely among species. In a survey of 33 species it was found that the longest chromosome in the complement varied from 22um, for x=9 species to 7um for the x=8 species. The average chromosome length for x=9 species is 14u, for x=7 species is 13um and in x=8 species is 9.2um.

Thus largest chromosomes are found in x=7 and x=9 species, the shortest in x=8 species.

DNA content has a very wide range of variation in Allium species which was measured for 65 species. The highest content is found in tetraploid A. globosum, to the lowest in diploid A. sibiricum (x=8).

There is no regular pattern either in relation to basic chromosome number grouping. x=7 species have the highest DNA content, the x=9 species are widely dispersed throughout the whole range of this measurement and x=8 species have the lowest content, but spreads from 48.6 to 15.2 picograms (2C), a difference equivalent from 6x to 2x! The highest amount among diploids is 51.2 picograms of DNA in the x=10 species A. grande.

In conclusion there is a massive variation in DNA content in Allium, independent of chromosome number and ploidy and we know very little why. In any case, the Allium genomes are huge genomes, even for those species at the lower end of the range, with 5 pc/1C nucleous.

As far as DNA content and species relationship, this information is not helpful. Presence of excessive amount of DNA does not reflect the presence of additional unique genes or of duplication of structural genes. Extra DNA is mostly highly repetitive DNA forming part of heterochromatic knobs interspersed along the chromosomes, telomeres and centromeres and rDNA regions. Amount of rDNA within x=8 species shows a two fold amount of variation, from 0.05% in A. sativum, to 0.121% in A. acutiflorum. The latter is a diploid with 5 pairs of nucleolar organizing chromosomes, yielding a maximum of 10 nucleoli in interphase. In situ hybridization with fluorescent probes makes possible detail identification of NOR chromosomes (Ricroch et al. 1992).


Onion:A. cepa 2n=2x=16 , genome size os over 16 gigabasepairs/1C, similar to hexaploid wheat. And 6x larger than maize. Very low gene density, one gene/168 kb. Littered by tracks of degenerated retroviral and transposons, covering ~50% of genome (Jaksie et al 2008) .

Center of origin: Primary: Northwestern India to Russia.

Secondary center: Iran, Turkey and the Mediterranean region.

Onions were domesticated in the Near East, Central Asia and Afghanistan and have been used since at least 5000 years ago. References to Allium species as food, medicine or religious objects can be traced back to the first Egyptian Dynasty, 3200 BC. Most likely it was introduced to Egypt much earlier.

No wild forms of A. cepa are found in nature.

Horticultural Groups:

1) Typicum, single bulbs, common onion.

2) Ascalonicum, shallot. Perennial onion, bulbs form clusters on the surface of the soil

3) Aggregatum: multiplier onions, potato onions. It grows closely packed clusters of bulbs underground rather than in the surface as ascalonicum types. It does not produce seed.

4) Proliferum/viviparum: small bulb, Top tree onion, bulbil propagation. These most likely are natural hybrids between A. cepa and A. fistulosum, based on karyotype studies by C banding). Normally the satellite chromosomes of the two species are distinguishable (Schubert et al 1983)

5) A. fistulosum or bunching onion, related species to A. cepa. Perennial, edible tops.

Wild relatives of onion:

A. oschaninii

A. pskemense

A. galanthum

A. roylei

a. altaicum

All these species have 2n=16.

A. altaicum might be ancestral to A. fistulosum

Hybridization:

A. cepa x A. oschaninii , roylei

Hybrids have reduced fertility :

A. cepa x galanthum

A. cepa x pskemense

Reduced chromosome pairing in the hybrids, due to the presence of inversions resulting in bridges and fragments.

Others hybrids are sterile.

A. fistulosum x A. cepa hybrids:

This hybrid may occur in nature. For example the species A. wakegi, which is similar to top onion or proliferum type, and is grown in Japan has been suggested that it is a hybrid between A. cepa (ascalonicum) and A. fistulosum. It is propagated vegetatively by inflorescence bulbils, since it does not produce seeds. Pollen fertility is restored in tetraploids by chromosome doubling. Some tetraploidized plants of A. wakegi, however are sterile, perhaps due to genic sterility. When tetraploid A. wakegi is crossed to diploid A. ascalonicum and A. fistulosum, 8 II + 8I chromosomal association is found indicating homology of each of the parental genomes. Furthermore, tetraploid A. wakegi x synthetic A. fistulosum x A. cepa allotetraploid hybrid is fully fertile, displaying 16 II in meiosis.

This hybrid has been obtained in several instances. One of the earliest attempts resulted in spontaneous chromosome doubling in the hybrid, originating a new cultivar called Beltsville Bunching Onion. The purpose of the hybridization was to transfer thrip resistance from A. fistulosum to A. cepa, but the hybrid look desirable as a new variety of bunching onion.

This type of hybrid is of interest also because both parental species differ on chiasma behavior. A. fistulosum has localized chiasmata; it always forms close to the centromere (proximal chiamata), while in A. cepa they forms anywhere in the chromosome (randomized chiasmata).

Differences in distribution of chiasmata exists in the genus. Some species has highly localized chiasmata. In this case, in addition to proximal chiasmata, localization can be distal or interstitial in other species.

The hybrid is perennial and non bulbing like the fistulosum parent.

The chiasmata are randomized in the hybrid like in the A. cepa parent. It has only 10% pollen fertility.

BC to A.fistulosum had some individuals with localized, others with randomized chiasmata and the majority with both types of chiasmata. Therefore, this trait might be controlled by the chromosomes themselves.

Allotetraploid and derivatives: Chromosome doubling the F1 hybrid results in completely fertile, true breeding, and regular meiosis with 16 II. Half of the bivalents have localized chiasmata, while the other half randomized chiasmata. When backcrossed to each parental diploid, triploids are obtained with the following chromosomal association: 8 II + 8I

Peffley et al. (1985) produced A.cepa-fistulosum addition lines by backcrossing the triploid to A. cepa.

Their objective was to transfer pink root resistance (Pyrenochaeta terrestris) to onion.

They observed possible recombination between the chromosomes of the two species in derived diploids from the additions. They displayed the leaf characteristic and pink root resistance of fistulosum. Hou and Peffley (2000) have now confirmed recombination, using FISH. Most of the recombinant plants were fertile.

Yaguchi et al 2008,09 developed a complete set of alien addition lines A. fistulosum-cepa (aggregatum). Four lines had higher polyphenol content. A phenylalanine ammonialyase gene was located on chromosome 2. Other genes in this family might be present in the other 3 addition lines, chromosomes 5, 6 and 8.

A. roylei has been used as a bridging species to introgress A. fistulosum genes into A. cepa (Khrustaleva and Kik, 2000). Floral biology: The onion flowers are perfect but protandrus, the stigma is receptive 3 days after initiation of pollen release, thus resulting in outcrossing by flies and bees.

Male sterility:

Cytoplasmic male sterility was found by Jones, (1925) after observing a plant of the cv. Italian Red, with bulbils in the inflorescence. This classical work resulting in the broad application of cms in plants took place at UC Davis in our department.

The cms plants have a normal meiosis, but the tetrads degenerate due to tapetum breakdown.

It has the typical [S]msms system, however, a small amount of pollen is produced at high temperatures.

Most onion varieties are heterozygous for the nuclear alleles (Msms)

Maintainer lines are [N] msms .

cms found later in other cvs.

N and S cytoplasms can be distinguished by RFLP analysis of chloroplast DNA. Insertion in [N] line chloroplasts DNA of 100 bp can be detected by amplification with primers by PCR (Havey 1995). This simplifies the identification of S lines. CMS is determined by mitochondria, however, since N and S have different alloplasmid origins, chlroplasts DNA also differs. Most likely the S cytoplasm was introduced by an interspecific cross of onion and an unknown species as female parent.

A second source of cms (T-cytoplasm) has been found in French cultivar `Jeune paille des Vertus'. Three independent genes, A, B and C restore fertility, which make it difficult to use this system. RFLP of cpDNA and mtDNA of N, S and T cytoplasms distinguish them by fragment profiles. N and T might derive from the same cytoplasm, but not N and S. These two cytoplasms can be distinguished also by Southern blots by hybridization of digested DNA to certain mitochondrial genes used as probes (Satoh et al. 1993)

Nuclear male sterile reported in A. fistulosum (jaja)

Male sterility is used for F1 hybrid production. These have superior seedling vigor and uniformity. Inbreeding depression is a problem when developing parental homozygous lines.

Important traits:

Bulb color in onions is determined by flavonoids.

Complex trait, determined by at least 5 genes (El Shafie and Davis 1967)

II color inhibitor

Ii incomplete dominance for inhibitor

ii allows color expression

C color factor

c colorless

G golden yellow

g brown

R-L- complementary action red/pink

These are the main genotypes:

iiCCGGRRLL red (anthocyanidin)

iiC-G-R-ll (or rrL-)pink

iiC-ggrrll chartreuse: (between yellow and green)

iiC-G-rrll yellow (dark to light) (only Quercitin)

iicc------ white recessive

II-------- white dominant

Ii-------- cream

Kim et al (2004) cloned most of the genes in the onion flavonoid pathway. They found that yellow color is due to a mutation in the gene DFR (dehydroflavonol 4-reductase). It corresponds to the R locus. However, this is a gene family with multiple copies which might contribute to intensity of red color. Additionally dominant alleles at a second locus, L2, linked to ANS (anthocyanidin synthase gene) at 6.3 cM could be involved in red bulb color. ANS corresponds to the L locus (Kim et al 2004;05;Jackse and Havey 2008). A new pink color was reported by Kim et al,. Pink color is recessive and undesirable. Pink recessiveness is due to low transcription of the gene (ANS). Kim et al (2005) have found that this is due to a 390bp insertion in the promoter explaining the molecular basis of the pink allele. They developed co-dominant markers for selection of red vs pink color. Markers allows to eliminate pink phenotypes when crossing red by white/yellow onions.

No candidate genes have been identified for locus G and its action in the flavonoid pathway is unknown.

C must be involved early in the pathways as a regulator. I also must be a regulator of the whole pathway.

Bulb color seems to be related to disease resistance; red onions are resistant to Botrytis, due to the presence of procatechuic ac. and catechol.

Recessive white can be distinguished from dominant white by subjecting bulb scales to NH4. Flavone responsible for color is present in the dominant genotype, so the scales turn yellow when exposed to the gas. II plants produce the color flavone, but the gene I inhibits its expression.

 

Day Length :

Cultivars classified according to photoperiod:

Short day onions: will bulb under short day conditions. Planted in the fall to harvest in the spring.

Long day onions: will bulb under long days. Planted in the spring, and harvested in the fall.

Processing vs Fresh market: Hi dry matter: fructan is the predominant carbohydrate, low dry matter: larger amounts of glucose, fructose and sucrose.

Flavor: It is produced mostly by cleavage of cysteine sulfoxide compounds mediated by allinase and soluble solids. These compounds derive from the amino acid cysteine. The sulfoxide compounds are odorless precursors to onion flavor and to lacrymatory compounds, which are volatile.

sulfoxides + water = thiosulfonates + pyruvate + ammonia

Pungency: There are mild, intermediate and pungent types. It is varietal but also affected by environment. There are genetic differences but not clear cut genetic studies. Processors prefer pungent varieties, whereas mild varieties are dedicated to fresh market. Low pungency is dominant over high pungency. Release of pyruvic acid is used to measure pungency. Pyruvate forms when the tissue is crushed and is highly correlated to flavor strength.

When the tissue is damaged, the precursors, which are odorless react with the enzyme alliinase to release the sulfenic acids plus ammonia and pyruvate. One of these is thiopropanal S-oxide (formed via propenyl sulfenic acid), which is the main lacrymatory principle in onions. The enzyme is in the vacuole, whereas the precursors are in the cytosol. So, when boiling a whole onion without damaging the tissue it becomes tasteless, because the enzyme is destroyed. (Garlic has allyl sulfenic acid, which does not lead to lacrymatory compounds).

Some of these compounds have medicinal properties by promoting cardiovascular health. This action might be based on the prevention of clot causing aggregation of platelets. Also they seem to be antiviral, to alleviate diabetes, asthma, cancer. Further some of them are probably potent antifungals. Varieties with higher S content and pungency have higher antiplatelet activity than mild onions.

Joint effort by scientists in New Zealand and Japan to produce tear-free onions by silencing the gene that produces the lachrymatory factor synthase preventing production of 1-propenyl containing thiosulfinates from 1-propenyl sulfenic acid. Called EverMild onion. (see Eady et al 2008)

Soluble solids: Determined by sugar content, which is important for the dehydration industry. A range from 5 to 16% can be found.

Diseases:

Downy mildew: (Peronospora destructor), genes s1 and s2 have been reported for resistance

Pink rot: (Pyrenochaeta terrestris), A. fistulosum is resistant

Virus. Yellow dwarf, A. fistulosum is immune.

Markers:

Isozymes have been used for the identification C. cepa-fistulosum of addition lines. Three of the 10 chromosome additions were identified by the enzymes: Pgm-1, Adh-1, Idh-1 and Pgi-1.

DNA based markers: Due to the fact that onions has one of the largest genomes of cultivated plants, 18 pc/1C, it is difficult to develop RFLPs. Only 4% of cDNA probes disclose polymorphism. A similar situation is found for RAPD primers, the majority fails to disclose polymorphism. Nevertheless, Havey et al. (1996) have constructed a low density linkage map of 12 groups based on 66 loci. This map have been expanded to 112 loci, including the male sterility gene ms, red bulb color gene and for the enzyme alliinase. 21% of the probes disclose duplicated loci, half of which are duplicated in tandem was commonly found (King et al. 1998). These may be the result of transpositional and retrotranspositional duplication of specific chromosome regions followed by rearrangements. Homology to retrotransposon sequences have been detected in intergenic spacers. More recently, van Heusden et al. (2000) have constructed an extensive AFLP map consisting of 692 markers in an interspecific F2 population obtained by crossing A. roylei x A. cepa. An alliinase and a downy mildew resistance gene are included in this map. EST markers added to this map by McCallum et al (2001) using SSCP . It includes 2 carbohydrate metabolism genes a a sulfurylase gene.

Khrustaleva et al (2005) have initiated the integration of genetic and physical maps using genomic insitu hybridization (GISH). Major gene for fructan content mapped by

Markers have been used in Allium for identification of interspecific hybrids and phylogenetic relationships.

Transformation: Onion is recalcitrant to transformation. Before only transient transformation with particle bombardment and Agrobacterium delivery system have been reported in onion. However, Eady et al (2000) reports successful A. tumefaciens mediated transformation using embryos as a source of explants.

Other cultivated species (x=8):

A. schoenoprassum: Chives, leaf onion, leaves are used as condiments. v. variable species, widespread through out the world. It is found even in the arctic, perennial, fertile.

Diploid, triploid and tetraploid forms are found.

A. chinense: Rakkyo, pickling vegetable in China , Rakkyo, also found as diploid, triploid and tetraploids. Tetraploids are the cultivated ones.

A. tuberosum, Chinese chives, grown for green leaves.

A. ampeloprasum, found as diploid, 3x, 4x, 5x, 6x and 8x

a) Garden leek is tetraploid, also known as A. porrum, 2n=4x=32

b) Kurrat, A. kurrat 4x short stature, very similar to leek, grown only for its leaves.

c) Great headed garlic, (2n=6x, 8x=48, 64), used as substitute for garlic, much larger, forms small bulbils in inflorescence. It is also known as Elephant garlic.

According to phylogenetic study of Hirschegger et al 2010 based on cpDNA, and rDN ITS, garden leek and kurrat closely related, but distant from great headed garlic. They seem to have different origins, being the 4x species autotetraploids and that 6x and 8x species allopolyploids. They propose to rename leek and kurrat as A. porrum.

Allium sativum: garlic

Onion and health

Wide range of health attributes such as antibiotic effects, antiasthmatic, anticarcinogenic, antiplatelet therefore diminishing stroke risk. (Griffith et al 2002). Frequent consumption of onion and garlic is inversely related to risk of common cancers in So European populations and China (Galeone et al 2008).



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Last modified, May 6, 2010

Carlos F Quiros, 1998