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is the genetic term for any complex of DNA and protein found in a nucleus of a cell |
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are the separate pieces of chromatin that behave as a unit during cell division |
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1/3 DNA, 1/3 histones, and 1/3 nonhistone proteins |
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DNA INTERACTIONS W/ HISTONES AND NONHISTONE PROTEINS |
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produces sufficient level of compaction to fit into a cell nucleus |
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small, positively charged, highly conserved - bind to & neutralize (-) charged DNA - 5 types-> H1,H2A,H2B,H3, & H4 -core histones make up the nucleosome |
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200-200000 molecules of each kind |
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LARGE VARIETY OF FUNCTION |
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structural role-> chromosome scaffold chromosome replicaiton -> DNA polymerase chromosome segregation->kinetochore protein active in transcription |
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DIFFERENT LEVELS OF CHROMOSOME COMPACTION |
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when stretched out, the DNA in a single cell would be about 6 ft long compaction allows the DNA to fit in a cell nucleus |
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a fundamental unit of chromosomal packing resemble beads on a string |
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SPACING AND STRUCTURE OF NUCLEOSOME AFFECT GENETIC FUNCTION |
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-accessibility for proteins that initiate transcription, replication, and further comp. -arrangement along chromatin is highly defined & varies in diff cell types &under diff conditions |
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THE NUCLEOSOME IS AN OCTAMER OF 2 EACH OF HISTONES H2A,H2B,H3, &H4 |
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160 bp of DNA wraps 2x around a nucleosome core 40bp linker DNA connects adj. nucleosomes histone H1 associates w/linker DNA as it enters & leaves nuclesome core |
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XRAY CRYSTALLOGRAPHY OF NUCLEOSOME |
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-DNA bends sharply at several places as it wraps around the core histone ocatmer - base seq. dictates preferred nucleosome potisions along the DNA |
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model of higher-order packaging 100 A nucleosomal chromatin is compacted in to 300A fiber by supercoiling |
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model for higher level of compaction several nonhistone proteins bind to chromatin every 60-100 kb and tether the 300A fiber into structure loops |
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may further condense chromosomes into a compact bundle for mitosis |
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KARYOTYPE OF A HUMAN FEMALE EXAMINED BY HIGH RESOLUTION G BANDING |
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-metaphase chromosomes stained w/ giemsa have alternating bands of light and dark stain -each contains many loops 1-10 mb long -banding patterns on each chromosome are highly reproducible |
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LOCATIONS OF GENE IN RELATION TO CHROMOSOMAL BANDS |
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short arm = p arm long arm = q arm w/in each arm, light and dark bands numbered |
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FLUORESCENT IN SITU HYBRIDIZATION (FISH) |
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depends on hybridization btwn metaphase chromosomes and a labeled DNA sequence |
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spread on a glass slide and denatured to make them single stranded |
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is labeled w/ fluorescent tag to make a probe |
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hybridizes to chromosomes at complementary regions |
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- is a variation of FISH - probes specific for each chromosome are labeled w/ different fluorescent dye |
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-highly condensed, usually inactive transcriptionally -darkly stained regions of chromosomes |
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condensed in all cells - most of the Y chromosome and all pericentromeric regions |
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condensed in only some cells and relaxed in others - position effect variegation, x chromosome in female mammals |
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relaxed, usually active transcriptionally - lightly stained region |
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changes in chromatin structure |
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of inactive genes are hidden in nucleosomes |
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transcription factors bind to enhancers and recruit chromatin remodeling proteins |
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exposed by removing or repositioning nucleosomes |
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white+ (w+) is normally located in euchromatin |
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can result in w+ gene being located adjacent to heterochromatin |
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-can be caused by spreading of heterochromatin into nearby gene -spreading can occur over > 1000 kb of chromatin - heterochromatin spreads further in some cells than in others |
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were used to identify genes involved in chromatin modification |
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-looked for changes in the amount of position effect variegation -mutations that enhanced heterochromatin formation made eyes more white -mutations that suppressed heterochromatin formation make less white eyes |
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SOME GENES ENCODE PROTEINS |
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that localize to heterochromatin |
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-can be modified w/ chemical groups -tails extend outward from nucleosome |
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ENZYMES CAN ADD CHEMICAL GROUPS |
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methyl group, phosphate group, ubiquitin modified tails can alter nucleosomes and bind chromatin modifier proteins |
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-histone acetyltransferases add acetyl group to histone tails -prevent close packing -favor expression of gene in euchromatin -reversed by histone deacetylases |
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-histone methyltransferases add methyl groups - affect depends on specific amino acid mod. -adding to H3 lysine 9 favors heterochromatin - reversed by methyltransferases |
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X-CHROMOSOME INACTIVATION |
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-in female mammals through heterochromatin formation -ex of facultative heterochromatin -x-linked genes in XX & XY express same level -random inactivation except x chromosome -Barr body |
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-early embryos bot X chromosomes are active -in humans random x inactivation 2 wks after fertilization -some maternal inactivation and some paternal adult females are mosaic X-linked |
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X-INACTIVATION IS INITIATED |
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by expression of the Xist gene |
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-x inactivation specific transcription -expressed on the inactive x, but not the active x |
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-is large, non-coding, cis-acting regulatory RNA -binds to the x-chromosome which expressed -initiates histone modification that result in heterochromatin formation |
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in human cells is 50 nt/sec |
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-800 hrs to replicate the human genome if there was only one origin of replication -most mammalian cells have 10000 origins -many origins are active at same time -accessible regions of DNA devoid in nucleosomes |
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DNA being replicated in both directions |
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disassembled and reformed during DNA replication |
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in nucleosomes w/in minutes of synthesis |
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associates w/DNA followed by 2 dimers of H2A and H2B |
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are composed of recycled and new histones |
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open to histone modification just after replication |
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that protect the ends of eukaryotic chromosmoes |
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-specific repetitive sequences and dont contain genes -Species-specific -TTAGGG in humans -> tetrahymena -250-1500 repeats w/ variable # btwn diff cells |
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REPLICATION AT ENDS OF CHROMOSOMES |
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-RNA primers used during DNA replication are removed, leaving a unreplicated DNA at 5' end -w/o a special mechanism, DNA would be lost from every new DNA strand at each cell |
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ARTIFICIAL CHROMOSOMES REQUIRE 3 KEY ELEMENTS |
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centromeres telomeres origins of replication |
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CONSEQUENCES OF DELETING KEY ELEMENTS |
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-no centromere no telomere - replicate segregation errors - no telomere - replicate degrade if linear |
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