Photographic display of total chromosome complement in a cell
"Idiogram" redirects here. Not to be confused with
ideogram.
A karyotype is the general appearance of the complete set of
chromosomes in the cells of a
species or in an individual organism, mainly including their sizes, numbers, and shapes.[1][2]Karyotyping is the process by which a karyotype is discerned by determining the chromosome complement of an individual, including the number of chromosomes and any abnormalities.
A karyogram or idiogram is a graphical depiction of a karyotype, wherein chromosomes are generally organized in pairs, ordered by size and position of centromere for chromosomes of the same size. Karyotyping generally combines
light microscopy and
photography in the
metaphase of the
cell cycle, and results in a
photomicrographic (or simply micrographic) karyogram. In contrast, a
schematic karyogram is a designed graphic representation of a karyotype. In schematic karyograms, just one of the sister
chromatids of each chromosome is generally shown for brevity, and in reality they are generally so close together that they look as one on photomicrographs as well unless the resolution is high enough to distinguish them. The study of whole sets of chromosomes is sometimes known as karyology.
Karyotypes describe the
chromosome count of an organism and what these chromosomes look like under a light
microscope. Attention is paid to their length, the position of the
centromeres, banding pattern, any differences between the
sex chromosomes, and any other physical characteristics.[3] The preparation and study of karyotypes is part of
cytogenetics.
The basic number of chromosomes in the
somatic cells of an individual or a species is called the somatic number and is designated 2n. In the
germ-line (the sex cells) the chromosome number is n (humans: n = 23).[4][5]p28 Thus, in humans 2n = 46.
So, in normal
diploid organisms,
autosomal chromosomes are present in two copies. There may, or may not, be
sex chromosomes.
Polyploid cells have multiple copies of chromosomes and
haploid cells have single copies.
The study of karyotypes is made possible by
staining. Usually, a suitable
dye, such as
Giemsa,[8] is applied after
cells have been arrested during
cell division by a solution of
colchicine usually in
metaphase or
prometaphase when most condensed. In order for the
Giemsa stain to adhere correctly, all chromosomal proteins must be digested and removed. For humans,
white blood cells are used most frequently because they are easily induced to divide and grow in
tissue culture.[9] Sometimes observations may be made on non-dividing (
interphase) cells. The sex of an unborn
fetus can be predicted by observation of interphase cells (see
amniotic centesis and
Barr body).
Observations
Six different characteristics of karyotypes are usually observed and compared:[10]
Differences in absolute sizes of chromosomes. Chromosomes can vary in absolute size by as much as twenty-fold between genera of the same family. For example, the legumes Lotus tenuis and Vicia faba each have six pairs of chromosomes, yet V. faba chromosomes are many times larger. These differences probably reflect different amounts of DNA duplication.
Differences in the position of
centromeres. These differences probably came about through
translocations.
Differences in relative size of chromosomes. These differences probably arose from segmental interchange of unequal lengths.
Differences in basic number of chromosomes. These differences could have resulted from successive unequal translocations which removed all the essential genetic material from a chromosome, permitting its loss without penalty to the organism (the dislocation hypothesis) or through fusion. Humans have one pair fewer chromosomes than the great apes. Human chromosome 2 appears to have resulted from the fusion of two ancestral chromosomes, and many of the genes of those two original chromosomes have been translocated to other chromosomes.
Differences in number and position of satellites.
Satellites are small bodies attached to a chromosome by a thin thread.
Differences in degree and distribution of
GC content (
Guanine-
Cytosine pairs versus
Adenine-
Thymine). In metaphase where the karyotype is typically studied, all DNA is condensed, but most of the time, DNA with a high GC content is usually less condensed, that is, it tends to appear as
euchromatin rather than
heterochromatin. GC rich DNA tends to contain more
coding DNA and be more
transcriptionally active.[11] GC rich DNA is lighter on
Giemsa staining.[12] Euchromatin regions contain larger amounts of
Guanine-
Cytosine pairs (that is, it has a higher
GC content). The staining technique using
Giemsa staining is called
G banding and therefore produces the typical "G-Bands".[12]
A full account of a karyotype may therefore include the number, type, shape and banding of the chromosomes, as well as other cytogenetic information.
Both the micrographic and schematic karyograms shown in this section have a standard chromosome layout, and display darker and lighter regions as seen on
G banding, which is the appearance of the chromosomes after treatment with
trypsin (to partially digest the chromosomes) and
staining with
Giemsa stain. Compared to darker regions, the lighter regions are generally more
transcriptionally active, with a greater ratio of
coding DNA versus
non-coding DNA, and a higher
GC content.[11]
Both the micrographic and schematic karyograms show the normal human
diploid karyotype, which is the typical composition of the
genome within a normal cell of the human body, and which contains 22 pairs of
autosomal chromosomes and one pair of
sex chromosomes (allosomes). A major exception to diploidy in humans is
gametes (sperm and egg cells) which are haploid with 23 unpaired chromosomes, and this
ploidy is not shown in these karyograms. The micrographic karyogram is converted to
grayscale, whereas the schematic karyogram shows the purple hue as typically seen on Giemsa stain (and is a result of its azure B component, which stains DNA purple).[14]
The schematic karyogram in this section is a graphical representation of the idealized karyotype. For each chromosome pair, the scale to the left shows the length in terms of million
base pairs, and the scale to the right shows the designations of the
bands and sub-bands. Such bands and sub-bands are used by the
International System for Human Cytogenomic Nomenclature to describe locations of
chromosome abnormalities. Each row of chromosomes is vertically aligned at
centromere level.
Human chromosome groups
Based on the karyogram characteristics of size, position of the
centromere and sometimes the presence of a
chromosomal satellite (a segment distal to a
secondary constriction), the human chromosomes are classified into the following groups:[15]
Very small, acrocentric (and 21, 22 with
satellite)
Alternatively, the human genome can be classified as follows, based on pairing, sex differences, as well as location within the
cell nucleus versus inside
mitochondria:
22
homologousautosomal chromosome pairs (chromosomes 1 to 22). Homologous means that they have the same genes in the same loci, and autosomal means that they are not sex chromomes.
Two
sex chromosome (in green rectangle at bottom right in the schematic karyogram, with adjacent silhouettes of typical representative
phenotypes): The most common karyotypes for
females contain two
X chromosomes and are denoted 46,XX;
males usually have both an X and a
Y chromosome denoted 46,XY. However, approximately 0.018% percent of humans are
intersex, sometimes due to variations in sex chromosomes.[16]
The
human mitochondrial genome (shown at bottom left in the schematic karyogram, to scale compared to the nuclear DNA in terms of
base pairs), although this is not included in micrographic karyograms in clinical practice. Its genome is relatively tiny compared to the rest.
Copy number
Schematic karyograms generally display a DNA copy number corresponding to the
G0 phase of the cellular state (outside of the replicative
cell cycle) which is the most common state of cells. The schematic karyogram in this section also shows this state. In this state (as well as during the G1 phase of the
cell cycle), each cell has 2 autosomal chromosomes of each kind (designated 2n), where each chromosome has one copy of each
locus, making a total copy number of 2 for each locus (2c). At top center in the schematic karyogram, it also shows the chromosome 3 pair after having undergone
DNA synthesis, occurring in the
S phase (annotated as S) of the cell cycle. This interval includes the
G2 phase and
metaphase (annotated as "Meta."). During this interval, there is still 2n, but each chromosome will have 2 copies of each locus, wherein each
sister chromatid (chromosome arm) is connected at the centromere, for a total of 4c.[17] The chromosomes on micrographic karyograms are in this state as well, because they are generally micrographed in metaphase, but during this phase the two copies of each chromosome are so close to each other that they appear as one unless the image resolution is high enough to distinguish them. In reality, during the G0 and G1 phases, nuclear DNA is dispersed as
chromatin and does not show visually distinguishable chromosomes even on micrography.
The copy number of the
human mitochondrial genome per human cell varies from 0 (erythrocytes)[18] up to 1,500,000 (
oocytes), mainly depending on the number of mitochondria per cell.[19]
Diversity and evolution of karyotypes
Although the
replication and
transcription of
DNA is highly standardized in
eukaryotes, the same cannot be said for their karyotypes, which are highly variable. There is variation between species in chromosome number, and in detailed organization, despite their construction from the same
macromolecules. This variation provides the basis for a range of studies in evolutionary
cytology.
In some cases there is even significant variation within species. In a review, Godfrey and Masters conclude:
In our view, it is unlikely that one process or the other can independently account for the wide range of karyotype structures that are observed ... But, used in conjunction with other phylogenetic data, karyotypic fissioning may help to explain dramatic differences in diploid numbers between closely related species, which were previously inexplicable.[20]
Although much is known about karyotypes at the descriptive level, and it is clear that changes in karyotype organization has had effects on the evolutionary course of many species, it is quite unclear what the general significance might be.
We have a very poor understanding of the causes of karyotype evolution, despite many careful investigations ... the general significance of karyotype evolution is obscure.
Instead of the usual gene repression, some organisms go in for large-scale elimination of
heterochromatin, or other kinds of visible adjustment to the karyotype.
Chromosome elimination. In some species, as in many
sciarid flies, entire chromosomes are eliminated during development.[22]
Chromatin diminution (founding father:
Theodor Boveri). In this process, found in some
copepods and
roundworms such as Ascaris suum, portions of the chromosomes are cast away in particular cells. This process is a carefully organised genome rearrangement where new telomeres are constructed and certain heterochromatin regions are lost.[23][24] In A. suum, all the somatic cell precursors undergo chromatin diminution.[25]
X-inactivation. The inactivation of one X chromosome takes place during the early development of mammals (see
Barr body and
dosage compensation). In
placental mammals, the inactivation is random as between the two Xs; thus the mammalian female is a mosaic in respect of her X chromosomes. In
marsupials it is always the paternal X which is inactivated. In human females some 15% of somatic cells escape inactivation,[26] and the number of genes affected on the inactivated X chromosome varies between cells: in
fibroblast cells up about 25% of genes on the Barr body escape inactivation.[27]
Number of chromosomes in a set
A spectacular example of variability between closely related species is the
muntjac, which was investigated by
Kurt Benirschke and
Doris Wurster. The diploid number of the Chinese muntjac, Muntiacus reevesi, was found to be 46, all
telocentric. When they looked at the karyotype of the closely related Indian muntjac, Muntiacus muntjak, they were astonished to find it had female = 6, male = 7 chromosomes.[28]
They simply could not believe what they saw ... They kept quiet for two or three years because they thought something was wrong with their tissue culture ... But when they obtained a couple more specimens they confirmed [their findings].
The number of chromosomes in the karyotype between (relatively) unrelated species is hugely variable. The low record is held by the
nematodeParascaris univalens, where the
haploid n = 1; and an ant: Myrmecia pilosula.[30] The high record would be somewhere amongst the
ferns, with the adder's tongue fern Ophioglossum ahead with an average of 1262 chromosomes.[31] Top score for animals might be the
shortnose sturgeonAcipenser brevirostrum at 372 chromosomes.[32] The existence of supernumerary or
B chromosomes means that chromosome number can vary even within one interbreeding population; and
aneuploids are another example, though in this case they would not be regarded as normal members of the population.
Fundamental number
The fundamental number, FN, of a karyotype is the number of visible major chromosomal arms per set of chromosomes.[33][34] Thus, FN ≤ 2 x 2n, the difference depending on the number of chromosomes considered single-armed (
acrocentric or
telocentric) present. Humans have FN = 82,[35] due to the presence of five acrocentric chromosome pairs:
13,
14,
15,
21, and
22 (the human
Y chromosome is also acrocentric). The fundamental autosomal number or autosomal fundamental number, FNa[36] or AN,[37] of a karyotype is the number of visible major chromosomal arms per set of
autosomes (non-
sex-linked chromosomes).
Ploidy
Ploidy is the number of complete sets of chromosomes in a cell.
Polyploidy, where there are more than two sets of homologous chromosomes in the cells, occurs mainly in plants. It has been of major significance in plant evolution according to
Stebbins.[38][39][40][41] The proportion of flowering plants which are polyploid was estimated by Stebbins to be 30–35%, but in grasses the average is much higher, about 70%.[42] Polyploidy in lower plants (
ferns,
horsetails and
psilotales) is also common, and some species of ferns have reached levels of polyploidy far in excess of the highest levels known in flowering plants. Polyploidy in animals is much less common, but it has been significant in some groups.[43]
Polyploid series in related species which consist entirely of multiples of a single basic number are known as
euploid.
Endopolyploidy occurs when in adult
differentiated tissues the cells have ceased to divide by
mitosis, but the
nuclei contain more than the original
somatic number of
chromosomes.[44] In the endocycle (
endomitosis or
endoreduplication) chromosomes in a 'resting' nucleus undergo
reduplication, the daughter chromosomes separating from each other inside an intactnuclear membrane.[45] In many instances, endopolyploid nuclei contain tens of thousands of chromosomes (which cannot be exactly counted). The cells do not always contain exact multiples (powers of two), which is why the simple definition 'an increase in the number of chromosome sets caused by replication without cell division' is not quite accurate. This process (especially studied in insects and some higher plants such as maize) may be a developmental strategy for increasing the productivity of tissues which are highly active in biosynthesis.[46] The phenomenon occurs sporadically throughout the
eukaryote kingdom from
protozoa to humans; it is diverse and complex, and serves
differentiation and
morphogenesis in many ways.[47]
See
palaeopolyploidy for the investigation of ancient karyotype duplications.
Aneuploidy
Aneuploidy is the condition in which the chromosome number in the cells is not the typical number for the species. This would give rise to a
chromosome abnormality such as an extra chromosome or one or more chromosomes lost. Abnormalities in chromosome number usually cause a defect in development.
Down syndrome and
Turner syndrome are examples of this.
Aneuploidy may also occur within a group of closely related species. Classic examples in plants are the genus Crepis, where the gametic (= haploid) numbers form the series x = 3, 4, 5, 6, and 7; and Crocus, where every number from x = 3 to x = 15 is represented by at least one species. Evidence of various kinds shows that trends of evolution have gone in different directions in different groups.[48] In primates, the
great apes have 24x2 chromosomes whereas humans have 23x2.
Human chromosome 2 was formed by a merger of ancestral chromosomes, reducing the number.[49]
Chromosomal polymorphism
Some species are
polymorphic for different chromosome structural forms.[50] The structural variation may be associated with different numbers of chromosomes in different individuals, which occurs in the ladybird beetle Chilocorus stigma, some
mantids of the genus Ameles,[51] the European shrew Sorex araneus.[52] There is some evidence from the case of the
molluscThais lapillus (the
dog whelk) on the
Brittany coast, that the two chromosome morphs are
adapted to different habitats.[53]
Species trees
The detailed study of chromosome banding in insects with
polytene chromosomes can reveal relationships between closely related species: the classic example is the study of chromosome banding in
Hawaiian drosophilids by
Hampton L. Carson.
In about 6,500 sq mi (17,000 km2), the
Hawaiian Islands have the most diverse collection of drosophilid flies in the world, living from
rainforests to
subalpine meadows. These roughly 800 Hawaiian drosophilid species are usually assigned to two genera, Drosophila and Scaptomyza, in the family
Drosophilidae.
The polytene banding of the 'picture wing' group, the best-studied group of Hawaiian drosophilids, enabled Carson to work out the evolutionary tree long before genome analysis was practicable. In a sense, gene arrangements are visible in the banding patterns of each chromosome. Chromosome rearrangements, especially
inversions, make it possible to see which species are closely related.
The results are clear. The inversions, when plotted in tree form (and independent of all other information), show a clear "flow" of species from older to newer islands. There are also cases of colonization back to older islands, and skipping of islands, but these are much less frequent. Using
K-Ar dating, the present islands date from 0.4 million years ago (mya) (
Mauna Kea) to 10mya (
Necker). The oldest member of the Hawaiian archipelago still above the sea is
Kure Atoll, which can be dated to 30 mya. The archipelago itself (produced by the
Pacific plate moving over a
hot spot) has existed for far longer, at least into the
Cretaceous. Previous islands now beneath the sea (
guyots) form the
Emperor Seamount Chain.[54]
All of the native Drosophila and Scaptomyza species in Hawaiʻi have apparently descended from a single ancestral species that colonized the islands, probably 20 million years ago. The subsequent
adaptive radiation was spurred by a lack of
competition and a wide variety of
niches. Although it would be possible for a single
gravid female to colonise an island, it is more likely to have been a group from the same species.[55][56][57][58]
There are other animals and plants on the Hawaiian archipelago which have undergone similar, if less spectacular, adaptive radiations.[59][60]
Chromosome banding
Chromosomes display a banded pattern when treated with some stains. Bands are alternating light and dark stripes that appear along the lengths of chromosomes. Unique banding patterns are used to identify chromosomes and to diagnose chromosomal aberrations, including chromosome breakage, loss, duplication, translocation or inverted segments. A range of different chromosome treatments produce a range of banding patterns: G-bands, R-bands, C-bands, Q-bands, T-bands and NOR-bands.
Depiction of karyotypes
Types of banding
Cytogenetics employs several techniques to visualize different aspects of chromosomes:[9]
G-banding is obtained with
Giemsa stain following digestion of chromosomes with
trypsin. It yields a series of lightly and darkly stained bands — the dark regions tend to be heterochromatic, late-replicating and AT rich. The light regions tend to be euchromatic, early-replicating and GC rich. This method will normally produce 300–400 bands in a normal,
human genome. It is the most common chromosome banding method.[61]
R-banding is the reverse of G-banding (the R stands for "reverse"). The dark regions are euchromatic (guanine-cytosine rich regions) and the bright regions are heterochromatic (thymine-adenine rich regions).
C-banding: Giemsa binds to
constitutive heterochromatin, so it stains
centromeres. The name is derived from centromeric or constitutive heterochromatin. The preparations undergo alkaline denaturation prior to staining leading to an almost complete depurination of the DNA. After washing the probe the remaining DNA is renatured again and stained with Giemsa solution consisting of methylene azure, methylene violet, methylene blue, and eosin. Heterochromatin binds a lot of the dye, while the rest of the chromosomes absorb only little of it. The C-bonding proved to be especially well-suited for the characterization of plant chromosomes.
Q-banding is a
fluorescent pattern obtained using
quinacrine for staining. The pattern of bands is very similar to that seen in G-banding. They can be recognized by a yellow fluorescence of differing intensity. Most part of the stained DNA is heterochromatin. Quinacrin (atebrin) binds both regions rich in AT and in GC, but only the AT-quinacrin-complex fluoresces. Since regions rich in AT are more common in heterochromatin than in euchromatin, these regions are labelled preferentially. The different intensities of the single bands mirror the different contents of AT. Other fluorochromes like DAPI or Hoechst 33258 lead also to characteristic, reproducible patterns. Each of them produces its specific pattern. In other words: the properties of the bonds and the specificity of the fluorochromes are not exclusively based on their affinity to regions rich in AT. Rather, the distribution of AT and the association of AT with other molecules like histones, for example, influences the binding properties of the fluorochromes.
Silver staining:
Silver nitrate stains the
nucleolar organization region-associated protein. This yields a dark region where the silver is deposited, denoting the activity of rRNA genes within the NOR.
Classic karyotype cytogenetics
In the "classic" (depicted) karyotype, a
dye, often
Giemsa(G-banding), less frequently
mepacrine (quinacrine), is used to stain bands on the chromosomes. Giemsa is specific for the
phosphate groups of
DNA. Quinacrine binds to the
adenine-
thymine-rich regions. Each chromosome has a characteristic banding pattern that helps to identify them; both chromosomes in a pair will have the same banding pattern.
Karyotypes are arranged with the short arm of the chromosome on top, and the long arm on the bottom. Some karyotypes call the short and long arms p and q, respectively. In addition, the differently stained regions and sub-regions are given numerical designations from
proximal to
distal on the chromosome arms. For example,
Cri du chat syndrome involves a deletion on the short arm of chromosome 5. It is written as 46,XX,5p-. The critical region for this syndrome is deletion of p15.2 (the
locus on the chromosome), which is written as 46,XX,del(5)(p15.2).[62]
Multicolor FISH (mFISH) and spectral karyotype (SKY technique)
Multicolor
FISH and the older spectral karyotyping are molecular
cytogenetic techniques used to simultaneously visualize all the pairs of
chromosomes in an organism in different colors.
Fluorescently labeled probes for each chromosome are made by labeling chromosome-specific DNA with different
fluorophores. Because there are a limited number of spectrally distinct fluorophores, a combinatorial labeling method is used to generate many different colors. Fluorophore combinations are captured and analyzed by a fluorescence microscope using up to 7 narrow-banded fluorescence filters or, in the case of spectral karyotyping, by using an
interferometer attached to a fluorescence microscope. In the case of an mFISH image, every combination of fluorochromes from the resulting original images is replaced by a
pseudo color in a dedicated image analysis software. Thus, chromosomes or chromosome sections can be visualized and identified, allowing for the analysis of chromosomal rearrangements.[63]
In the case of spectral karyotyping, image processing software assigns a
pseudo color to each spectrally different combination, allowing the visualization of the individually colored chromosomes.[64]
Multicolor FISH is used to identify structural chromosome aberrations in cancer cells and other disease conditions when Giemsa banding or other techniques are not accurate enough.
Digital karyotyping
Digital karyotyping is a technique used to quantify the DNA copy number on a genomic scale. Short sequences of DNA from specific loci all over the genome are isolated and enumerated.[65] This method is also known as
virtual karyotyping. Using this technique, it is possible to detect small alterations in the human genome, that cannot be detected through methods employing metaphase chromosomes. Some loci deletions are known to be related to the development of cancer. Such deletions are found through digital karyotyping using the loci associated with cancer development.[66]
Chromosome abnormalities can be numerical, as in the presence of extra or missing chromosomes, or structural, as in
derivative chromosome,
translocations,
inversions, large-scale deletions or duplications. Numerical abnormalities, also known as
aneuploidy, often occur as a result of
nondisjunction during
meiosis in the formation of a
gamete;
trisomies, in which three copies of a chromosome are present instead of the usual two, are common numerical abnormalities. Structural abnormalities often arise from errors in
homologous recombination. Both types of abnormalities can occur in gametes and therefore will be present in all cells of an affected person's body, or they can occur during
mitosis and give rise to a
genetic mosaic individual who has some normal and some abnormal cells.
In humans
Chromosomal abnormalities that lead to disease in humans include
Turner syndrome results from a single X chromosome (45,X or 45,X0).
Klinefelter syndrome, the most common male chromosomal disease, otherwise known as 47,XXY, is caused by an extra X chromosome.
Trisomy 9, believed to be the 4th most common trisomy, has many long lived affected individuals but only in a form other than a full trisomy, such as trisomy 9p syndrome or mosaic trisomy 9. They often function quite well, but tend to have trouble with speech.
Also documented are trisomy 8 and trisomy 16, although they generally do not survive to birth.
Some disorders arise from loss of just a piece of one chromosome, including
Cri du chat (cry of the cat), from a truncated short arm on chromosome 5. The name comes from the babies' distinctive cry, caused by abnormal formation of the larynx.
Angelman syndrome – 50% of cases have a segment of the long arm of chromosome 15 missing; a deletion of the maternal genes, example of
imprinting disorder.
Prader-Willi syndrome – 50% of cases have a segment of the long arm of chromosome 15 missing; a deletion of the paternal genes, example of imprinting disorder.
The next stage took place after the development of genetics in the early 20th century, when it was appreciated that chromosomes (that can be observed by karyotype) were the carrier of genes. The term karyotype as defined by the
phenotypic appearance of the
somatic chromosomes, in contrast to their
genic contents was introduced by
Grigory Levitsky who worked with Lev Delaunay,
Sergei Navashin, and
Nikolai Vavilov.[67][68][69][70] The subsequent history of the concept can be followed in the works of
C. D. Darlington[71] and
Michael JD White.[4][13]
Investigation into the human karyotype took many years to settle the most basic question: how many chromosomes does a normal
diploid human cell contain?[72] In 1912,
Hans von Winiwarter reported 47 chromosomes in
spermatogonia and 48 in
oogonia, concluding an
XX/XOsex determination mechanism.[73]Painter in 1922 was not certain whether the diploid of humans was 46 or 48, at first favoring 46,[74] but revised his opinion from 46 to 48, and he correctly insisted on humans having an
XX/XY system.[75] Considering the techniques of the time, these results were remarkable.
Joe Hin Tjio working in
Albert Levan's lab[76] found the chromosome count to be 46 using new techniques available at the time:
Squashing the preparation on the slide forcing the chromosomes into a single plane
Cutting up a photomicrograph and arranging the result into an indisputable karyogram.
The work took place in 1955, and was published in 1956. The karyotype of humans includes only 46 chromosomes.[77][29] The other
great apes have 48 chromosomes.
Human chromosome 2 is now known to be a result of an end-to-end fusion of two ancestral ape chromosomes.[78][79]
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