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free120/Block 1/Question#9 (reveal difficulty score)
During an experiment, a Southern blot ...
Gene rearrangement πŸ” / πŸ“Ί / 🌳
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 +9 
submitted by βˆ—waterloo(118),
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oYu acn kool ta tihs iqonsetu atlageicyls.rt

  1. hSuenort btlo &-;-tg os AND.
  2. yeth rae igsun an enoeaclnesdu ot rrsetafn btis, shti dtos'en ryeall amne yhitangn ot uoy .tey
  3. DANc si pibngro rof lgeins ncont-iaimsobomtnugluln n.ioreg

'Thtas apm,ntoirt sceaeub wno oyu nowk tyhe ear lnokgoi at hte tnsncato nigreo fo munilogominblu aak Fc fo het yevha ain.hc

oS wyh is hist cDNA htta oedecns Fc, hgligint up lony neco ni hreto ituesss, but hte smae DcNA glstih pu iulpetlm miest ni elna 2 noe(b rmo?)rwa Tsih is sca,beeu ta eth nboe orrwam omndar toarcnmbeinio of iltgh ro yeavh ihnca rc,ocsu dan ruo DAcN is tesnrep in ftinrfdee ardmon ibtoncisoanm tihw lghti i.hasnc Ttah escicifp AcND nceqeeus teyh sedu dntd'i ngaehc nbeeewt teh itesssu or nvee wihint eth bnoe mwor,ra adn ti will noly ndbi ot the one eneg seuqnce.e Teh tacf s'it tignilhg up lielmtpu etsim ellst uoy ttha the amse ensueqec si pen,trse oyru tiqesoun si why so mayn tesim.

lsoA a dies ,teno htunores btslo aer meti m,sngicnou os bsal sue heotr mhtosde ot od eth msae ntghi e(ikl PR)C, tub ehsuonrt sltbo ear isllt het tebs hnew it ecsmo to cecihkng iumimnboognlul nda T elcl tpreeocr ngee angaeertrermn.




 +6 
submitted by βˆ—sugaplum(453),
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hiTs qoesitnu si sgikan buota VDJ nemtarregnaer whchi epnhpas in hte nobe a.omwrr eTh egens ear lla hpdpeco pu aeucsbe the B llec is yinrtg to eartgene a qieuun imnnoaiotcb rfo sti rcertpeo
emspil nt.ecpocs.. dod wnrgiod

prCeaht 3 of "how the inmemu smeyst rswk"o - weaomes oobk

varunmehru  in the question stem, they are asking about a constant region. VDJ rearrangement is for the variable. It doesn't make sense :( +4  
sallz  Both the constant (heavy chains) and the light chains undergo gene rearrangement. The heavy chain undergoes V(D)J random recombinations, while the light chain undergo VJ random recombinations. So gene rearrangement could work for both regions. +13  
azibird  The constant region does not undergo recombination. That's why it's called constant. It's just right next to the variable region though, so they get expressed together as one protein. That's why the constant-labeled DNA region is variable length here. +4  
chaosawaits  You're right that the constant region is not undergoing recombination. The multiple bands are not do to constant region rearrangement; it's due to the variable chain rearranging and then the cDNA probe that is specific to the constant region attaches to each of these rearrangements, since it hasn't changed +  



 +1 
submitted by βˆ—em_goldman(29),
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I vlbieee lps( eorcrct em fi 'Im or)nwg oyu oludw vhae riliasm enhtourS otbl sstleur esen ni B csell nouenidggr icsatom uioeyntrmta,hp btu thta keast caelp in the ycadsenor lipmodhy suties, not in hte ebon wa.morr




 +1 
submitted by βˆ—azibird(227),
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I rade this tuiqsoen as nalinyzag inbouolugsimmln ni eht boen rrwmoa vs otu irgtnciaclu l.hlypripaere I nwo rzlieea isth is rwg,on and tyhe rea sutj ynaagizln the DNA fo rtieeffnd ellc li,appnuoto klei a apyehectot rfo axlep,em otn ougnmimuisoblln rpesetn ni het e.vrli

iWht ihts ldrcaee up, I nca ees who V)(JD irmcotioanneb oclud emka het iliagonr DNA eoscnti r.eosrth uBt owh olcdu (V)JD tnaoebmnroiic psolybsi aekm eht ADN ictoesn egonr?l ehreT era dnasb on the lge ttah are hrgihe clrueloam hgwiet ni hte bneo arw.mro ceoboimanRint lyon treshson teh ,ADN mneoose eeaspl ccrotre .em

Aosl why is teehr on dnab ni the beno rwmrao thta si teh mase ezsi sa elrripheap a?bsnd yeluSr the CsHS ehav ont enurngeod etconn,riimoba rehwe is hiter D??NA?

azibird  Here is the figure from Kaplan Immunology. See how recombination only makes the sequence shorter? https://imgur.com/a/fI2jHfu Kaplan also says: "While heavy chain gene segments are undergoing recombination, the enzyme terminal deoxyribonucleotidyl transferase (Tdt) randomly inserts bases (without a template on the complementary strand) at the junctions of V, D, and J segments (N-nucleotide addition). The random addition of the nucleo- tide generates junctional diversity." However, this would not account for the equal stepwise lengthening of DNA, which is clearly from these recombinable units. +1  



 +1 
submitted by βˆ—hello(398),
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Eoincgh 'loroaews@t ec.tmomn

ihsT Q si alstlsieeny gaiskn "htaW is the inocutfn of hturSeon bt"?lso

nArews: ehT ofuticnn fo Shterou tbols is to kehcc for BRC ro TRC neeg egetemrnrranas.

eSe lr@ewa'oots nxeltonapia for htwa si iifsapcellcy nggio no sa fra sa yhw tereh rea erifndtef bndsa ta detirfefn .peacls A,gian teh awrsen ot siht si "gnee "treemrnnrgaea but tloewro@a lasxniep ni wtih oerm .ateidl




 +0 
submitted by βˆ—benwhite_dotcom(653),
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Stuherno oslbt rea cmonlmyo sude in laoominigmcul ,ssetdiu as het onuhster olbt oaswll rfo het ydsut fo ADN i.totaernlsa thWa is lramlnyo neo egne giocfarinontu ealterd to iummen siblnloug in otms seussti edttaseromns ptulmlie itrdnfefe bsand ni the obne a,wormr itdaicniev of eeng .mearntreaergn ihTs is islyaacbl how ew tcreae enw o.tebsnadii iReetacv csesopesr aer loyacplnol liutpem(l )sandb; aemuikel, in atsnctr,o si oloclnnoam (legsin badn.)

ali  I still don’t understand this one. Could you provide a better explanation? +1  
benwhite_dotcom  The cDNA tag is tagging a constant region common to immunoglobulins, so it normally only finds the one band corresponding to that particular gene (the bands travel different amounts due to their differing size/weight). In the bone marrow sample, that gene has rearranged itself, so the cDNA clone instead tags multiple different genes that are of different sizes on the gel (each one has that same constant region the cDNA is tagging, but with different stuff around it such that the restriction enzyme has cut it up differently). I’d be happy for someone to step in and do a better job on that explanation. +19  
em_goldman  A Southern blot starts by cutting DNA strands at a particular (short) site and running them through gel electrophoresis, so identical DNA sequences get cut at the same site and thus are the same length, so they are at the same place on the gel. If there's lots of different sequences, the restriction endonuclease (the scissors) cut the DNA at different places, leading to strands that were the same length originally but are now lots of different lengths -> different places on the gel. But how do you know this is the same gene, just with different mutations? The Southern blot uses a probe to look for a more specific (long) region of DNA that you know is in the target gene. So even though there are mutations causing the less-specific endonuclease to cut the DNA at different parts, the overall architecture of the gene is similar enough that the probe can bind, thus we know it's the same gene. (And in bone marrow WBCs, the mechanism here is genetic rearrangement.) +2  



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