Editing Junk DNA

{{Short description|Those parts of a genome with no function}}
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'''Junk DNA''' ('''non-functional DNA''') is a DNA sequence that has no known biological function.<ref name="Eddy2012">{{cite journal | vauthors = Eddy SR | title = The C-value paradox, junk DNA and ENCODE | journal = Current Biology | volume = 22 | issue = 21 | pages = R898–R899 | date = November 2012 | pmid = 23137679 | doi = 10.1016/j.cub.2012.10.002 | bibcode = 2012CBio...22.R898E | s2cid = 28289437 | doi-access = free | author-link = Sean Eddy }}</ref><ref name="PalazzoGregory2014">{{cite journal | vauthors = Palazzo AF, Gregory TR | title = The case for junk DNA | journal = PLOS Genetics | volume = 10 | issue = 5 | pages = e1004351 | date = May 2014 | pmid = 24809441 | pmc = 4014423 | doi = 10.1371/journal.pgen.1004351 | doi-access = free }}</ref> Most organisms have some junk DNA in their [[genome]]s&mdash;mostly [[pseudogene]]s and fragments of [[transposons]] and viruses&mdash;but it is possible that some organisms have substantial amounts of junk DNA.<ref>{{cite journal | vauthors = Gil R, Latorre A | title = Factors behind junk DNA in bacteria | journal = Genes | volume = 3 | issue = 4 | pages = 634–650 | date = October 2012 | pmid = 24705080 | pmc = 3899985 | doi = 10.3390/genes3040634 | doi-access = free }}</ref>

All protein-coding regions are generally considered to be functional elements in genomes. Additionally, non-protein coding regions such as genes for ribosomal RNA and transfer RNA, regulatory sequences, origins of replication, centromeres, telomeres, and scaffold attachment regions are considered as functional elements. (See [[Non-coding DNA]] for more information.)

It is difficult to determine whether other regions of the genome are functional or nonfunctional. There is considerable controversy over which criteria should be used to identify function. Many scientists have an evolutionary view of the genome and they prefer criteria based on whether DNA sequences are preserved by natural selection.<ref name="Ohno1972b">{{cite journal | vauthors = Ohno S | date = 1972 | title = An argument for the genetic simplicity of man and other mammals | journal = Journal of Human Evolution | volume = 1 | issue = 6 | pages = 651–662 | doi = 10.1016/0047-2484(72)90011-5 | bibcode = 1972JHumE...1..651O }}</ref><ref name="Morange2014">{{cite journal | vauthors = Morange M | title = Genome as a multipurpose structure built by evolution | journal = Perspectives in Biology and Medicine | volume = 57 | issue = 1 | pages = 162–171 | date = 2014 | pmid = 25345709 | doi = 10.1353/pbm.2014.0008 | s2cid = 27613442 | url = https://hal.archives-ouvertes.fr/hal-01480552/file/ARTICLE%20ENCODE%20MM%2070114%20corrige%C2%A6%C3%BC.pdf }}</ref><ref name="Palazzo&Kejiou2022">{{cite journal | vauthors = Palazzo AF, Kejiou NS | title = Non-Darwinian Molecular Biology | journal = Frontiers in Genetics | volume = 13 | pages = 831068 | year = 2022 | pmid = 35251134 | pmc = 8888898 | doi = 10.3389/fgene.2022.831068 | doi-access = free }}</ref> Other scientists dispute this view or have different interpretations of the data.<ref name = Germain_et_al_2014>{{ cite journal | vauthors = Germain PL, Ratti E, and Boem F | date = 2014 | title = Junk or functional DNA? ENCODE and the function controversy | journal = Biology & Philosophy | volume = 29 | issue = 6 | pages = 807–821 | doi = 10.1007/s10539-014-9441-3| s2cid = 254277794 }}</ref><ref name = Mattick2023>{{ cite journal | last = Mattick | first = John S | date = 2023 | title =  RNA out of the mist | journal = Trends in Genetics | volume = 39 | issue = 3 | pages = 187–207 | doi = 10.1016/j.tig.2022.11.001 | pmid = 36528415 | s2cid = 254768457 }}</ref><ref name="kellis">{{cite journal |vauthors=Kellis M, Wold B, Snyder MP, Bernstein BE, Kundaje A, Marinov GK, Ward LD, Birney E, Crawford GE, Dekker J, Dunham I, Elnitski LL, Farnham PJ, Feingold EA, Gerstein M, Giddings MC, Gilbert DM, Gingeras TR, Green ED, Guigo R, Hubbard T, Kent J, Lieb JD, Myers RM, Pazin MJ, Ren B, Stamatoyannopoulos JA, Weng Z, White KP, Hardison RC |date=April 2014 |title=Defining functional DNA elements in the human genome |journal=Proceedings of the National Academy of Sciences of the United States of America|volume=111 |issue=17 |pages=6131–6138 |bibcode=2014PNAS..111.6131K |doi=10.1073/pnas.1318948111 |pmc=4035993 |pmid=24753594 |doi-access=free}}</ref>

== History ==

The idea that only a fraction of the human genome could be functional dates back to the late 1940s. The estimated mutation rate in humans suggested that if a  large fraction of those mutations were deleterious then the human species could not survive such a mutation load (genetic load). This led to predictions in the late 1940s by one of the founders of population genetics, [[J.B.S. Haldane]], and by Nobel laureate [[Hermann Joseph Muller|Hermann Muller]], that only a small percentage of the human genome contains functional DNA elements (genes) that can be destroyed by mutation.<ref name = Muller1950 >{{cite journal | vauthors = Muller HJ | title = Our load of mutations | journal = American Journal of Human Genetics | volume = 2 | issue = 2 | pages = 111–176 | date = June 1950 | pmid = 14771033 | pmc = 1716299 }}</ref><ref name= Haldane1949>{{ cite journal | last = Haldane | first = JBS | date = 1949 | title = The rate of mutation of human genes | journal = Hereditas | volume = 35 | pages = 267–273 |  doi = 10.1111/j.1601-5223.1949.tb03339.x | doi-access = free }}</ref>  (see [[Genetic load]] for more information)

In 1966 Muller reviewed these predictions and concluded that the human genome could only contain about 30,000 genes based on the number of deleterious mutations that the species could tolerate.<ref name = Muller1966>{{cite journal | vauthors = Muller HJ | title = The gene material as the initiator and the organizing basis of life | journal = The American Naturalist | volume = 100 | issue = 915 | pages = 493-517 | date = September 1966 | doi = 10.1086/282445 | jstor = 2459205 }}</ref> Similar predictions were made by other leading experts in molecular evolution who concluded that the human genome could not contain more than 40,000 genes and that less than 10% of the genome was functional.<ref name = Kimura1968>{{cite journal | vauthors = Kimura M | title = Evolutionary rate at the molecular level | journal = Nature | volume = 217 | issue = 5129 | pages = 624–626 | date = February 1968 | pmid = 5637732 | doi = 10.1038/217624a0 | s2cid = 4161261 | bibcode = 1968Natur.217..624K }}</ref><ref name = KingJukes1969>{{cite journal | vauthors = King JL, Jukes TH | title = Non-Darwinian evolution | journal = Science | volume = 164 | issue = 3881 | pages = 788–798 | date = May 1969 | pmid = 5767777 | doi = 10.1126/science.164.3881.788 | bibcode = 1969Sci...164..788L }}</ref><ref name = Ohno1972b/><ref name = Ohta&Kimura1971>{{cite journal | vauthors = Ohta T, Kimura M | title = Functional organization of genetic material as a product of molecular evolution | journal = Nature | volume = 233 | issue = 5315 | pages = 118–119 | date = September 1971 | pmid = 16063236 | doi = 10.1038/233118a0 | s2cid = 13344748 | bibcode = 1971Natur.233..118O }}</ref>

The size of genomes in various species was known to vary considerably and there did not seem to be a correlation between genome size and the complexity of the species. Even closely related species could have very different genome sizes. This observation led to what came to be known as the [[C-value paradox]].<ref name = Thomas1971 >{{cite journal | vauthors = Thomas CA | title = The genetic organization of chromosomes | journal = Annual Review of Genetics | volume = 5 | pages = 237–256 | date = 1971 | pmid = 16097657 | doi = 10.1146/annurev.ge.05.120171.001321 }}</ref> The paradox was resolved with the discovery of repetitive DNA and the observation that most of the differences in genome size could be attributed to repetitive DNA.<ref name = Thomas1971/><ref name = Britten&Kohne1968 >{{cite journal | vauthors = Britten RJ, Kohne DE | title = Repeated sequences in DNA. Hundreds of thousands of copies of DNA sequences have been incorporated into the genomes of higher organisms | journal = Science | volume = 161 | issue = 3841 | pages = 529–540 | date = August 1968 | pmid = 4874239 | doi = 10.1126/science.161.3841.529 | bibcode = 1968Sci...161..529B }}</ref> Some scientists thought that most of the repetitive DNA was involved in regulating gene expression but many scientists thought that the excess repetitive DNA was nonfunctional.<ref name = Britten&Davidson1969 >{{cite journal | vauthors = Britten RJ, Davidson EH | title = Gene regulation for higher cells: a theory | journal = Science | volume = 165 | issue = 3891 | pages = 349–357 | date = July 1969 | pmid = 5789433 | doi = 10.1126/science.165.3891.349 | bibcode = 1969Sci...165..349B }}</ref><ref name = Thomas1971/><ref name = Gregory2005>{{cite book | last = Gregory | first = TR | date = 2005 | chapter = Genome Size Evolution in Animals | title = The Evolution of the Genome | pages = 3–87 | publisher = Elsevier }}</ref><ref name = Lewin1972>{{ cite book | last = Lewin | first = Benjamin | date = 1974 | chapter = Chapter 4: Sequences of Eukaryotic DNA | title = Gene Expression-2: Eukaryotic Chromosomes | publisher = John Wiley & Sons }}</ref><ref name=Lewin1974c>{{cite journal|last = Lewin | first = Benjamin | date = 1974 | title = Sequence Organization of Eukaryotic DNA: Defining the Unit of Gene Expression | journal = Cell | volume =  1 | issue = 3 | pages = 107–111 | doi = 10.1016/0092-8674(74)90125-1 }}</ref>

[[File:Tomoko Harada cropped 1 Tomoko Harada 201611.png|thumb|[[Tomoko Ohta]] (Tomoko Harada) developed the nearly neutral theory that led to an understanding of how slightly deleterious junk DNA could be maintained in the genomes of species with small effective population sizes. In 2015 she was awarded the [[Crafoord Prize]] by the Royal Swedish Academy (with Richard Lewontin).]]At about the same time (late 1960s) the newly developed technique of [[Cot analysis|C<sub>0</sub>t analysis]] was refined to include RNA:DNA hybridization leading to the discovery that considerably less than 10% of the human genome was complementary to mRNA and this DNA was in the unique (non-repetitive) fraction. This confirmed the predictions made from genetic load arguments and was consistent with the idea that much of the repetitive DNA is nonfunctional.<ref name = Lewin1972b>{{ cite book | last = Lewin | first = Benjamin | date = 1974 | chapter = Chapter 5: Transcription and Processing of RNA | title = Gene Expression-2: Eukaryotic Chromosomes | publisher = John Wiley & Sons }}</ref><ref name = OBrian1973>{{cite journal | vauthors = O'Brien SJ | title = On estimating functional gene number in eukaryotes | journal = Nature | volume = 242 | issue = 115 | pages = 52–54 | date = March 1973 | pmid = 4512011 | doi = 10.1038/newbio242052a0 }}</ref><ref name = Bishop1974>{{cite journal | vauthors = Bishop JO | title = The gene numbers game | journal = Cell | volume = 2 | issue = 2 | pages = 81–86 | date = June 1974 | pmid = 4616752 | doi = 10.1016/0092-8674(74)90095-6 }}</ref>

The idea that large amounts of eukaryotic genomes could be nonfunctional conflicted with the prevailing view of evolution in 1968 since it seemed likely that nonfunctional DNA would be eliminated by natural selection. The development of the [[Neutral theory of molecular evolution|neutral theory]] and the [[Nearly neutral theory of molecular evolution|nearly neutral theory]] provided a way out of this problem since it allowed for the preservation of slightly deleterious nonfunctional DNA in accordance with fundamental principles of population genetics.<ref name=KingJukes1969/><ref name=Kimura1968/><ref name = Kimura&Ohta1971>{{cite journal | vauthors = Kimura M, Ohta T | title = Protein polymorphism as a phase of molecular evolution | journal = Nature | volume = 229 | issue = 5285 | pages = 467–469 | date = February 1971 | pmid = 4925204 | doi = 10.1038/229467a0 | s2cid = 4290427 | bibcode = 1971Natur.229..467K }}</ref>

The term "junk DNA" began to be used in the late 1950s<ref>{{cite thesis |type=MA |last=Sweet |first=Amalia |date=2022 |title=Requiem for a Gene: The Problem of Junk DNA for the Molecular Paradigm |publisher=University of Chicago| url = https://knowledge.uchicago.edu/record/5164}}</ref> but [[Susumu Ohno]] popularized the term in a 1972 paper titled "So much 'junk' DNA in our genome"<ref name=Ohno1972a>{{cite journal | vauthors = Ohno S | title = So much "junk" DNA in our genome | journal = Brookhaven Symposia in Biology | volume = 23 | pages = 366–370 | date = 1972 | pmid = 5065367 }}</ref> where he summarized the current evidence that had accumulated by then.<ref name = Ohno1972a/> In a second paper that same year, he concluded that 90% of mammalian genomes consisted of nonfunctional DNA.<ref name = Ohno1972b/> The case for junk DNA was summarized in a lengthy paper by David Comings in 1972 where he listed four reasons for proposing junk DNA:<ref name="Comings1972a">{{cite book |title=Advances in human genetics |vauthors=Comings DE |date=1972 |publisher=Springer |pages=237–431 |chapter=The structure and function of chromatin}}</ref>

# some organisms have a lot more DNA than they seem to require (C-value [[paradox]]),
# current estimates of the number of genes (in 1972) are much less than the number that can be accommodated,
# the mutation load would be too large if all the DNA were functional, and
# some junk DNA clearly exists.

The discovery of [[intron]]s in the 1970s seemed to confirm the views of junk DNA proponents because it meant that genes were very large and even huge genomes could not accommodate large numbers of genes. The proponents of junk DNA tended to dismiss intron sequences as mostly nonfunctional DNA (junk) but junk DNA opponents advanced a number of hypotheses attributing functions of various sort to intron sequences.<ref name="Morange2020a">{{cite book | last = Morange | first = Michel | date = 2020 | chapter = Chapter 17: Split Genes and Splicing | title = The Black Box of Biology: A History of the Molecular Revolution | publisher = Harvard University Press }}</ref><ref name="Gilbert1978">{{cite journal | vauthors = Gilbert W | title = Why genes in pieces? | journal = Nature | volume = 271 | issue = 5645 | pages = 501 | date = February 1978 | pmid = 622185 | doi = 10.1038/271501a0 | s2cid = 4216649 | doi-access = free | bibcode = 1978Natur.271..501G }}</ref><ref name="Gilbert1985">{{cite journal | vauthors = Gilbert W | title = Genes-in-pieces revisited | journal = Science | volume = 228 | issue = 4701 | pages = 823–824 | date = May 1985 | pmid = 4001923 | doi = 10.1126/science.4001923 | bibcode = 1985Sci...228..823G }}</ref><ref name="Crick1978">{{cite journal | vauthors = Crick F | title = Split genes and RNA splicing | journal = Science | volume = 204 | issue = 4390 | pages = 264–271 | date = April 1979 | pmid = 373120 | doi = 10.1126/science.373120 | bibcode = 1979Sci...204..264C }}</ref><ref name="Doolittle1978">{{cite journal | last = Doolittle | first = W.F. | date = 1978 |  title = Genes in pieces: were they ever together? | journal = Nature | volume =  272 | issue = 5654 | pages = 581–582 | doi = 10.1038/272581a0 | bibcode = 1978Natur.272..581D | s2cid = 4162765 | doi-access = free }}</ref>

[[File:Francis Crick crop.jpg|thumb|[[Francis Crick]] and others promoted the idea that transposons were examples of selfish DNA and were responsible for the proliferation of junk DNA.]]By 1980 it was apparent that most of the repetitive DNA in the human genome was related to [[transposons]]. This prompted a series of papers and letters describing transposons as selfish DNA that acted as a parasite in genomes and produced no fitness advantage for the organism.<ref name=Doolittle&Sapienza1980>{{cite journal | vauthors = Doolittle WF, Sapienza C | title = Selfish genes, the phenotype paradigm and genome evolution | journal = Nature | volume = 284 | issue = 5757 | pages = 601–603 | date = April 1980 | pmid = 6245369 | doi = 10.1038/284601a0 | s2cid = 4311366 | bibcode = 1980Natur.284..601D }}</ref><ref name = Orgel&Crick1980>{{cite journal | vauthors = Orgel LE, Crick FH | title = Selfish DNA: the ultimate parasite | journal = Nature | volume = 284 | issue = 5757 | pages = 604–607 | date = April 1980 | pmid = 7366731 | doi = 10.1038/284604a0 | s2cid = 4233826 | bibcode = 1980Natur.284..604O }}</ref><ref name=Dover1980>{{cite journal | vauthors = Dover G | title = Ignorant DNA? | journal = Nature | volume = 285 | issue = 5767 | pages = 618–620 | date = June 1980 | pmid = 7393318 | doi = 10.1038/285618a0 | s2cid = 4261755 | doi-access = free | bibcode = 1980Natur.285..618D }}</ref><ref name = Dover&Doolittle1980>{{cite journal | vauthors = Dover G, Doolittle WF | title = Modes of genome evolution | journal = Nature | volume = 288 | issue = 5792 | pages = 646–647 | date = December 1980 | pmid = 6256636 | doi = 10.1038/288646a0 | s2cid = 8938434 | doi-access = free | bibcode = 1980Natur.288..646D }}</ref><ref name = Jain1980>{{cite journal | vauthors = Jain HK | title = Incidental DNA | journal = Nature | volume = 288 | issue = 5792 | pages = 647–648 | date = December 1980 | pmid = 7453799 | doi = 10.1038/288647a0 | s2cid = 31899622 | doi-access = free | bibcode = 1980Natur.288..647J }}</ref>

Opponents of junk DNA interpreted these results as evidence that most of the genome is functional and they developed several hypotheses advocating that transposon sequences could benefit the organism or the species.<ref name = Cavalier-Smith1980>{{cite journal | vauthors = Cavalier-Smith T | title = How selfish is DNA? | journal = Nature | volume = 285 | issue = 5767 | pages = 617–618 | date = June 1980 | pmid = 7393317 | doi = 10.1038/285617a0 | s2cid = 27111068 | doi-access = free | bibcode = 1980Natur.285..617C }}</ref> The most important opponent of junk DNA at this time was [[Thomas Cavalier-Smith]] who argued that the extra DNA was required to increase the volume of the nucleus in order to promote more efficient transport across the nuclear membrane.<ref name = Cavalier-Smith1978>{{cite journal | vauthors = Cavalier-Smith T | title = Nuclear volume control by nucleoskeletal DNA, selection for cell volume and cell growth rate, and the solution of the DNA C-value paradox | journal = Journal of Cell Science | volume = 34 | pages = 247–278 | date = December 1978 | pmid = 372199 | doi = 10.1242/jcs.34.1.247 }}</ref>

The positions of the two sides of the controversy hardened with one side believing that evolution was consistent with large amounts of junk DNA and the other side believing that natural selection should eliminate junk DNA. These differing views of evolution were highlighted in a letter from [[Thomas H. Jukes|Thomas Jukes]], a proponent of junk DNA, to Francis Crick on December 20, 1979:<ref>{{ cite web | url = https://profiles.nlm.nih.gov/spotlight/sc/catalog/nlm:nlmuid-101584582X199-doc | title = letter to Francis Crick | first = Jukes | last = Thomas | date = December 29, 1979 | website = National Institutes of Health (USA) }}</ref>

<blockquote>"Dear Francis, I am sure that you realize how frightfully angry a lot of people will be if you say that much of the DNA is junk. The geneticists will be angry because they think that DNA is sacred. The Darwinian evolutionists will be outraged because they believe every change in DNA that is accepted in evolution is necessarily an adaptive change. To suggest anything else is an insult to the sacred memory of Darwin."</blockquote>

The other point of view was expressed by [[Roy John Britten]] and Kohne in their seminal paper on repetitive DNA.<ref name = Britten&Kohne1968/>

<blockquote>"A concept that is repugnant to us is that about half of the DNA of higher organisms is trivial or permanently inert (on an evolutionary timescale)."</blockquote>

==Junk DNA and non-coding DNA==

There is considerable confusion in the popular press and in the scientific literature about the distinction between non-coding DNA and junk DNA.

According to an article published in 2021 in American Scientist:

{{Blockquote |Close to 99 percent of our genome has been historically classified as noncoding, useless "junk" DNA. Consequently, these sequences were rarely studied.<ref>{{ cite journal | vauthors =  Mortola E, Long M | date = 2021 | title = Turning Junk into Us: How Genes Are Born | journal = American Scientist | volume = 109 | pages = 174–182 }}</ref> }}

A book published in 2020 states:

{{Blockquote | When it was first discovered, the nongenic DNA was sometimes called—somewhat derisively by people who did not know better—"junk DNA" because it had no obvious utility, and they foolishly assumed that if it was not carrying coding information it must be useless trash.<ref>{{ cite book | vauthors = McHughen A | date = 2020 | title = DNA Demystified: Unraveling the Double Helix | publisher = Oxford University Press | place = New York, New York, USA}}</ref> }}

The common theme is that the original proponents of junk DNA thought that all non-coding DNA was junk.<ref name="PalazzoGregory2014" /><ref name="Palazzo&Kejiou2022" /> This claim has been attributed to a paper by [[David Comings]] in 1972<ref name = Comings1972a /> where he is reported to have said that junk DNA refers to ''all'' non-coding DNA.<ref name="Gregory2005" /> But Comings never said that. In that paper he discusses non-coding genes for ribosomal RNA and tRNAs and non-coding regulatory DNA and he proposes several possible functions for the bulk of non-coding DNA.<ref name = Comings1972a/> In another publication from the same year Comings again discusses the term junk DNA with the clear understanding that it does not include non-coding regulatory sequences.<ref name = Comings1972b>{{ cite journal | last = Comings | first = DE | date = 1972 | title = Review of ''Evolution of Genetics Systems'' | journal = American Journal of Human Genetics | volume = 25 | pages = 340–342 }}</ref>

The idea that all non-coding DNA was thought to be junk has been criticized by numerous authors for distorting the history of junk DNA;<ref name="Eddy2012" /><ref name = Niu&Jiang2013>{{ cite journal | vauthors = Niu DK, Jiang L | date= 2013 | title =  Can ENCODE tell us how much junk DNA we carry in our genome? | journal =  Biochemical and Biophysical Research Communications | volume = 430 | issue= 4 | pages = 1340–1343 | doi = 10.1016/j.bbrc.2012.12.074| pmid= 23268340 }}</ref><ref name="Grauretal2013" /><ref>{{cite journal | vauthors = Graur D, Zheng Y, Azevedo RB | date = 2015 | title = An evolutionary classification of genomic function | journal = Genome Biology and Evolution | volume = 7 | issue = 3 | pages = 642–645 | doi = 10.1093/gbe/evv021| pmid = 25635041 | pmc = 5322545 }}</ref><ref name="PalazzoGregory2014"/> for example:
{{blockquote | It is simply not true that noncoding DNA has long been dismissed as worthless junk and that functional hypotheses have only recently been proposed - despite the frequency with which this cliché is repeated in media reports and in the introduction of far too many scientific studies.<ref name = Elliotetal2014>{{ cite journal | vauthors = Elliott TA, Linquist S, and Gregory TR | date = 2014 | title = Conceptual and empirical challenges of ascribing functions to transposable elements | journal = The American Naturalist | volume = 184 | issue = 1 |pages = 14–24 | doi = 10.1086/676588 | pmid = 24921597 | s2cid = 14549993 | url = http://philsci-archive.pitt.edu/11636/1/Conceptual_and_Empirical_Challenges_%28preprint_version%29.pdf }}</ref>}}
Some of the criticisms have been strong:
{{blockquote | Revisionist claims that equate noncoding DNA with junk merely reveal that people who are allowed to exhibit their logorrhea in Nature and other glam journals are as ignorant as the worst young-earth creationists.<ref name = Graur2017>{{cite book |last=Graur |first=Dan |date=2017 |editor-last=Saitou |editor-first=Naruya |title=Evolution of the Human Genome I |publisher=Springer |pages=19–60 |chapter=Rubbish DNA: The functionless fraction of the human genome }}</ref>}}

== Functional vs non-functional ==
The main challenge of identifying junk DNA is to distinguish between "functional" and "non-functional" sequences. This is non-trivial, but there is some good evidence for both categories.

=== Functional ===
[[Coding region|Protein-coding sequences]] are the most obvious functional sequences in genomes. However, they make up only 1-2% of most vertebrate genomes. However, there are also ''functional'' but ''non-coding'' DNA sequences<ref name="PalazzoGregory2014" /> such as [[regulatory sequence]]s, [[Origin of replication|origins of replication]], and [[centromere]]s.<ref name="Watson1965">{{cite book | vauthors = Watson J | title = Molecular Biology of the Gene | date = 1965 | publisher = W. A. Benjamin, Inc. | place = New York, New York, USA}}</ref> These sequences are usually conserved in evolution and make up another 3-8% of the human genome.<ref name=":0">{{cite journal | vauthors = Siepel A, Bejerano G, Pedersen JS, Hinrichs AS, Hou M, Rosenbloom K, Clawson H, Spieth J, Hillier LW, Richards S, Weinstock GM, Wilson RK, Gibbs RA, Kent WJ, Miller W, Haussler D | title = Evolutionarily conserved elements in vertebrate, insect, worm, and yeast genomes | journal = Genome Research | volume = 15 | issue = 8 | pages = 1034–1050 | date = August 2005 | pmid = 16024819 | pmc = 1182216 | doi = 10.1101/gr.3715005 }}</ref>

The Encyclopedia of DNA Elements ([[ENCODE]]) project reported that detectable biochemical activity was observed in regions covering at least 80% of the human genome, with biochemical activity defined primarily as being transcribed.<ref name="Nature489p57">{{cite journal | title = An integrated encyclopedia of DNA elements in the human genome | journal = Nature | volume = 489 | issue = 7414 | pages = 57–74 | date = September 2012 | pmid = 22955616 | pmc = 3439153 | doi = 10.1038/nature11247 | bibcode = 2012Natur.489...57T | collaboration = The ENCODE Project Consortium | vauthors = Dunham I, Kundaje A, Aldred SF, Collins PJ, Davis CA, Doyle F, etal }}</ref> While these findings were announced as the demise of junk DNA<ref name="Pennisi 2012">{{cite journal | vauthors = Pennisi E | title = Genomics. ENCODE project writes eulogy for junk DNA | journal = Science | volume = 337 | issue = 6099 | pages = 1159, 1161 | date = September 2012 | pmid = 22955811 | doi = 10.1126/science.337.6099.1159 }}</ref><ref name="Casane et al 2015">{{cite journal | vauthors = Casane D, Fumey J, Laurenti P | title = [ENCODE apophenia or a panglossian analysis of the human genome] | journal = Médecine/Sciences | volume = 31 | issue = 6–7 | pages = 680–686 | date = 2015 | pmid = 26152174 | doi = 10.1051/medsci/20153106023 }}</ref> it is important to point out that transcription does not mean a sequence is "functional", analogous to some meaningless text that can be transcribed or copied without having any meaning.<ref name="observer">{{cite news |date=February 24, 2013 |title=Scientists attacked over claim that 'junk DNA' is vital to life |work=The Observer |url=https://www.theguardian.com/science/2013/feb/24/scientists-attacked-over-junk-dna-claim |vauthors=McKie R}}</ref><ref name="Eddy2012" /><ref name="Eddy 2013">{{cite journal | vauthors = Eddy SR | title = The ENCODE project: missteps overshadowing a success | journal = Current Biology | volume = 23 | issue = 7 | pages = R259–R261 | date = April 2013 | pmid = 23578867 | doi = 10.1016/j.cub.2013.03.023 | doi-access = free | bibcode = 2013CBio...23.R259E }}</ref><ref name="doolittle2013">{{cite journal | vauthors = Doolittle WF | title = Is junk DNA bunk? A critique of ENCODE | journal = Proceedings of the National Academy of Sciences of the United States of America | volume = 110 | issue = 14 | pages = 5294–5300 | date = April 2013 | pmid = 23479647 | pmc = 3619371 | doi = 10.1073/pnas.1221376110 | bibcode = 2013PNAS..110.5294D | doi-access = free | author-link = W. Ford Doolittle }}</ref><ref name="Brunet and Doolittle 2014">{{cite journal | vauthors = Brunet TD, Doolittle WF | title = Getting "function" right | journal = Proceedings of the National Academy of Sciences of the United States of America | volume = 111 | issue = 33 | pages = E3365 | date = August 2014 | pmid = 25107292 | pmc = 4143013 | doi = 10.1073/pnas.1409762111 | doi-access = free | bibcode = 2014PNAS..111E3365P }}</ref><ref name="PalazzoGregory2014" /><ref name="Grauretal2013">{{cite journal | vauthors = Graur D, Zheng Y, Price N, Azevedo RB, Zufall RA, Elhaik E | title = On the immortality of television sets: "function" in the human genome according to the evolution-free gospel of ENCODE | journal = Genome Biology and Evolution | volume = 5 | issue = 3 | pages = 578–590 | year = 2013 | pmid = 23431001 | pmc = 3622293 | doi = 10.1093/gbe/evt028 }}</ref><ref name="Doolittle et al 2014">{{cite journal | vauthors = Doolittle WF, Brunet TD, Linquist S, Gregory TR | title = Distinguishing between "function" and "effect" in genome biology | journal = Genome Biology and Evolution | volume = 6 | issue = 5 | pages = 1234–1237 | date = May 2014 | pmid = 24814287 | pmc = 4041003 | doi = 10.1093/gbe/evu098 }}</ref><ref name="Morange2014" /><ref name="Niu&Jiang2013" /><ref name="kellis" />

=== Non-functional ===
Non-functional DNA is rare in [[bacterial genome]]s which typically have an extremely high gene density, with only a few percent being not protein-coding.<ref>{{cite journal | vauthors = Zhao Z, Cristian A, Rosen G | title = Keeping up with the genomes: efficient learning of our increasing knowledge of the tree of life | journal = BMC Bioinformatics | volume = 21 | issue = 1 | pages = 412 | date = September 2020 | pmid = 32957925 | pmc = 7507296 | doi = 10.1186/s12859-020-03744-7 | doi-access = free }}</ref>

However, in most animal or plant genomes, a large fraction of DNA is non-functional, given that there is no obvious selective pressure on these sequences. More importantly, there is strong evidence that these sequences are not functional in other ways (using the human genome as example):

(1) Repetitive elements, especially mobile elements make up a large fraction of the human genome, such as [[Retrotransposon#LTR retrotransposons|LTR retrotransposons]] (8.3% of total genome), [[Short interspersed nuclear element|SINEs]] (13.1% of total genome) including [[Alu elements]], [[Long interspersed nuclear element|LINEs]] (20.4% of total genome), SVAs (SINE-[[Variable number tandem repeat|VNTR]]-Alu) and [[Transposable element#Classification|Class II DNA transposons]] (2.9% of total genome).<ref>{{cite journal | vauthors = Treangen TJ, Salzberg SL | title = Repetitive DNA and next-generation sequencing: computational challenges and solutions | journal = Nature Reviews. Genetics | volume = 13 | issue = 1 | pages = 36–46 | date = November 2011 | pmid = 22124482 | pmc = 3324860 | doi = 10.1038/nrg3117 }}</ref> Many of these sequences are the descendents of ancient virus infections and are thus "non-functional" in terms of human genome function.

(2) Many sequences can be deleted as shown by comparing genomes. For instance, an analysis of 14,623 individuals identified 42,765 [[Structural variation|structural variants]] in the human genome of which 23.4% affected multiple genes (by deleting them or part of them). This study also found 47 deletions of >1 MB, showing that large chunks of the human genome can get deleted without obvious consequences.<ref>{{cite journal | vauthors = Abel HJ, Larson DE, Regier AA, Chiang C, Das I, Kanchi KL, Layer RM, Neale BM, Salerno WJ, Reeves C, Buyske S, Matise TC, Muzny DM, Zody MC, Lander ES, Dutcher SK, Stitziel NO, Hall IM | title = Mapping and characterization of structural variation in 17,795 human genomes | journal = Nature | volume = 583 | issue = 7814 | pages = 83–89 | date = July 2020 | pmid = 32460305 | pmc = 7547914 | doi = 10.1038/s41586-020-2371-0 | bibcode = 2020Natur.583...83A }}</ref>

(3) Only a small fraction of the human genome is conserved, indicating that there is no strong (functional) [[Evolutionary pressure|selection pressure]] on these sequences, so they can rather freely mutate.<ref name=":0" /><ref>{{cite book | vauthors = Graur D | date = 2016 | title = Molecular and Genome Evolution | publisher = Sinauer Associates, Inc. | place = Sunderland MA (USA) | isbn = 9781605354699}}</ref> About 11% or less of the human genome is conserved<ref name="Rands">{{cite journal | vauthors = Rands CM, Meader S, Ponting CP, Lunter G | title = 8.2% of the Human genome is constrained: variation in rates of turnover across functional element classes in the human lineage | journal = PLOS Genetics | volume = 10 | issue = 7 | pages = e1004525 | date = July 2014 | pmid = 25057982 | pmc = 4109858 | doi = 10.1371/journal.pgen.1004525 | doi-access = free }}</ref><ref name="Christmas et al 2023">{{cite journal | vauthors = Christmas MJ, Kaplow IM, Genereux DP, Dong MX, Hughes GM, Li X, Sullivan PF, Hindle AG, Andrews G, Armstrong JC, Bianchi M, Breit AM, Diekhans M, Fanter C, Foley NM, Goodman DB, Goodman L, Keough KC, Kirilenko B, Kowalczyk A, Lawless C, Lind AL, Meadows JR, Moreira LR, Redlich RW, Ryan L, Swofford R, Valenzuela A, Wagner F, Wallerman O, Brown AR, Damas J, Fan K, Gatesy J, Grimshaw J, Johnson J, Kozyrev SV, Lawler AJ, Marinescu VD, Morrill KM, Osmanski A, Paulat NS, Phan BN, Reilly SK, Schäffer DE, Steiner C, Supple MA, Wilder AP, Wirthlin ME, Xue JR, Birren BW, Gazal S, Hubley RM, Koepfli KP, Marques-Bonet T, Meyer WK, Nweeia M, Sabeti PC, Shapiro B, Smit AF, Springer MS, Teeling EC, Weng Z, Hiller M, Levesque DL, Lewin HA, Murphy WJ, Navarro A, Paten B, Pollard KS, Ray DA, Ruf I, Ryder OA, Pfenning AR, Lindblad-Toh K, Karlsson EK | title = Evolutionary constraint and innovation across hundreds of placental mammals | journal = Science | volume = 380 | issue = 6643 | pages = eabn3943 | date = April 2023 | pmid = 37104599 | pmc = 10250106 | doi = 10.1126/science.abn3943 | hdl-access = free | hdl = 10230/59591 }}</ref> and about 7% is under [[Negative selection (natural selection)|purifying selection]].<ref name="Halldorsonetal2022">{{cite journal | vauthors = Halldorsson BV, Eggertsson HP, Moore KH, Hauswedell H, Eiriksson O, Ulfarsson MO, Palsson G, Hardarson MT, Oddsson A, Jensson BO, Kristmundsdottir S, Sigurpalsdottir BD, Stefansson OA, Beyter D, Holley G, Tragante V, Gylfason A, Olason PI, Zink F, Asgeirsdottir M, Sverrisson ST, Sigurdsson B, Gudjonsson SA, Sigurdsson GT, Halldorsson GH, Sveinbjornsson G, Norland K, Styrkarsdottir U, Magnusdottir DN, Snorradottir S, Kristinsson K, Sobech E, Jonsson H, Geirsson AJ, Olafsson I, Jonsson P, Pedersen OB, Erikstrup C, Brunak S, Ostrowski SR, Thorleifsson G, Jonsson F, Melsted P, Jonsdottir I, Rafnar T, Holm H, Stefansson H, Saemundsdottir J, Gudbjartsson DF, Magnusson OT, Masson G, Thorsteinsdottir U, Helgason A, Jonsson H, Sulem P, Stefansson K | title = The sequences of 150,119 genomes in the UK Biobank | journal = Nature | volume = 607 | issue = 7920 | pages = 732–740 | date = July 2022 | pmid = 35859178 | pmc = 9329122 | doi = 10.1038/s41586-022-04965-x | hdl-access = free | bibcode = 2022Natur.607..732H | hdl = 20.500.11815/3726 }}</ref>

Opponents of junk DNA argue that biochemical activity detects functional regions of the genome that are not identified by sequence conservation or purifying selection.<ref name="Mattick&Dinger2013">{{cite journal | vauthors = Liu G, Mattick JS, Taft RJ | title = A meta-analysis of the genomic and transcriptomic composition of complex life | journal = Cell Cycle | volume = 12 | issue = 13 | pages = 2061–2072 | date = July 2013 | pmc = 4685169 | doi = 10.1186/1877-6566-7-2 | doi-access = free | pmid = 23759593 }}</ref><ref name="Mattick2023" /><ref name="Mattick 2023b">{{cite journal | vauthors = Mattick JS | title = A Kuhnian revolution in molecular biology: Most genes in complex organisms express regulatory RNAs | journal = BioEssays | volume = 45 | issue = 9 | pages = e2300080 | date = September 2023 | pmid = 37318305 | doi = 10.1002/bies.202300080 | doi-access = free }}</ref> According to some scientists, until a region in question has been shown to have additional features, beyond what is expected of the null hypothesis, it should provisionally be labelled as non-functional.<ref name="PalazzoLee2015">{{cite journal | vauthors = Palazzo AF, Lee ES | title = Non-coding RNA: what is functional and what is junk? | journal = Frontiers in Genetics | volume = 6 | pages = 2 | year = 2015 | pmid = 25674102 | pmc = 4306305 | doi = 10.3389/fgene.2015.00002 | doi-access = free }}</ref>

== See also ==
* [[ENCODE|ENCODE Project]]
* [[Human genome]]
* [[Comparative genomics]]
* [[Non-coding DNA]]
* [[Non-coding RNA]]

== References ==
{{Reflist}}

[[Category:DNA]]