C s phn t ca di truyn

Hnh 16.1 Cu trc ADN c xc nh nh th no? Cc khi nim chnh

16.1. ADN l vt cht di truyn 16.2. Nhiu protein phi hp vi nhau trong qu trnh sao chp v sa cha ADN 16.3. Mi nhim sc th gm mt phn t ADN c ng gi cng vi cc protein
Tng quan

Khi nim

ADN l vt cht di truyn

Ngy nay, ngay c hc sinh tiu hc cng c nghe ni v ADN, trong khi cc nh khoa hc thng xuyn thao tc vi ADN trong phng th nghim nhm lm thay i tnh trng di truyn ca cc t bo v c th. Tuy vy, n u th k XX, phn t no lm nhim v di truyn vn cn cha r v lc cu hi ny l mt trong nhng thch thc ln nht vi cc nh sinh hc.

16.1

Bn ch dn vn hnh s sng

o nm 1953, James Watson v Francis Crick gy chn ng cng ng khoa hc bng vic cng b m hnh chui xon kp v cu trc phn t ca axit deoxyribonucleic, c gi tt l ADN. Hnh 16.1 cho thy Watson (tri) v Crick ang say sa ngm nhn m hnh ADN c dng bng v hp v dy thp ca h. Tri qua hn 50 nm, m hnh ny xut thn ch l mt xut mi dn tr thnh biu tng ca sinh hc hin i. ADN, hp cht di truyn, chnh l phn t ng ngng m nht trong thi i ca chng ta. Trong thc t, cc yu t di truyn ca Mendel cng nh cc gen nm trn nhim sc th ca Morgan u cha ADN. Trn quan im ha hc, c th ni ti sn di truyn m mi ngi chng ta li cho th h sau chnh l cc phn t ADN c trn 46 nhim sc th m chng ta c tha hng t cc thn sinh (b, m) v ADN c trong cc ti th c truyn li theo dng m. Trong tt c cc phn t c trong t nhin, cc axit nucleic l c nht, v nh v kh nng t sao chp (ti bn) t cc n phn thnh phn. Trong thc t, c im con ci ging b, m l kt qu ca qu trnh sao chp chnh xc ADN v s di truyn ca n qua cc th h. Thng tin di truyn c m ha bng ngn ng ha hc ca ADN v c ti bn mi t bo trong c th ca mi ngi chng ta. Chnh ngn ng lp trnh ca ADN iu khin qu trnh pht trin cc tnh trng v ha sinh, gii phu, sinh l v mt mc nht nh l tp tnh mi c th sinh vt. Chng ny cp n vic bng cch no cc nh khoa hc chng minh c ADN l vt cht di truyn v bng cch no Watson v Crick pht hin ra cu trc phn t ca n. ng thi, chng ta cng s thy bng cch no ADN c th sao chp (c s phn t ca di truyn) v c sa cha. Cui cng, chng ta s kho st xem ADN cng vi protein ng gi nh th no trong nhim sc th.

Tm kim vt cht di truyn: Qu trnh tm hiu khoa hc

Khi nhm nghin cu ca T. H. Morgan cho thy cc gen nm dc theo cc nhim sc th (xem m t Chng 15), hai thnh phn ha hc ca nhim sc th - ADN v protein - c coi l hai hp cht ng vin cho vai tr vt cht di truyn. Cho n nhng nm 1940, cc bng chng "ng h" protein dng nh u th hn; c bit, mt s nh ha sinh xp protein vo nhm cc i phn t va c tnh c hiu chc nng cao va c tnh a dng vn l nhng yu cu thit yu ca vt cht di truyn. Ngoi ra, lc hiu bit v cc axit nucleic cn hn ch; dng nh cc thuc tnh vt l v ha hc ca chng khng tng ng vi s a dng phong ph ca cc tnh trng di truyn biu hin c th mi c th sinh vt khc nhau. Quan im ny sau c thay i dn t kt qu ca mt s nghin cu vi sinh vt. Ging nh cc nghin cu ca Mendel v Morgan, mt trong nhng yu t quan trng nht xc nh c tnh nht qun ca vt cht di truyn chnh l vic la chn c loi sinh vt th nghim ph hp. Vai tr di truyn ca ADN c pht hin u tin vi khun v virut; chng c c im n gin hn so vi u H lan, rui gim v ngi. phn tip theo ca chng ny, chng ta s ln theo qu trnh tm hiu khoa hc t cc nh khoa hc tm ra v xc nh c vai tr l vt cht di truyn ca ADN.

305

Bng chng l ADN c th bin i vi khun

Chng ta c th theo di qu trnh khm ph ra vai tr di truyn ca ADN ngc tr v nm 1928. Vo nm , mt y s qun y ngi Anh tn l Frederick Griffith, trong n lc tm kim vcxin phng bnh vim phi, tin hnh nghin cu vi khun Streptococcus pneumoniae l tc nhn gy bnh vim phi cc loi ng vt c v. Griffith c hai chng (ging) vi khun; mt chng c (gy bnh) v mt chng khng c
Hnh 16.2

Nghin cu pht hin

Tnh trng di truyn c th truyn gia cc chng vi khun khc nhau hay khng?
Th nghim Frederick Griffith nghin cu hai chng vi khun Streptococcus pneumoniae. Chng vi khun S (khun lc trn) gy vim phi chut; y l chng c v t bo ca chng c lp v khng c h thng bo v ng vt. Chng vi khun R (khun lc nhn) khng c lp v v khng c (khng gy bnh). th nghim qu trnh pht sinh bnh, Griffith tim hai chng vi khun vo chut th nghim nh s di y:
Cc t bo S sng (i chng) Cc t bo R sng (i chng) Cc t bo S cht bi nhit (i chng) Hn hp t bo S cht v R sng

(khng gy bnh). Griffith rt ngc nhin khi pht hin ra rng khi cc t bo ca chng c b dit bi nhit (un nng) khi c trn vi cc t bo sng ca chng khng c li c th sinh ra cc t bo con gy c (Hnh 16.2). Hn na, tnh trng tp nhim ny c di truyn cho tt c cc t bo vi khun th h con xut pht t t bo bin i ban u. R rng, mt cht ha hc no (lc cha r bn cht) ca cc t bo gy c cht gy nn s bin i di truyn ny. Griffith gi hin tng ny l bin np v c chng ta ngy nay nh ngha l qu trnh mt t bo tip nhn ADN t mi trng bn ngoi, dn n s thay i kiu gen v kiu hnh. Cng b ca Griffith m ng cho mt nghin cu c trin khai trong sut 14 nm sau bi mt nh virut hc ngi M tn l Oswald Avery nhm mc ch xc nh bn cht ca cht bin np. Avery tp trung vo ba nhm hp cht c nhiu kh nng hn c l ADN, ARN (mt loi axit nucleic khc) v protein. Avery tin hnh ph v t bo ca chng vi khun gy c cht bi un nng, ri tin hnh chit xut cc thnh phn t dch chit t bo. mi phng thc th nghim, Avery tin hnh x l lm bt hot tng nhm cht. Sau , dch chit sau khi x l c trn v kim tra kh nng bin np vo chng vi khun khng c cn sng. Kt qu th nghim cho thy ch khi ADN c duy tr (khng b bt hot) hin tng bin np m Griffith m t mi din ra. Nm 1944, Avery v cc ng nghip ca mnh l Maclyn McCarty v Colin MacLeod cng b rng: ADN chnh l cht bin np. Pht hin ny ca h c cho n bi nhiu ngi quan tm, nhng cng c khng t hoi nghi, mt phn c th bi v nhiu ngi quen vi t tng xem protein l vt cht di truyn ph hp hn. Hn na, nhiu nh khoa hc khng thuyt phc vi quan im cho rng cc gen ca vi khun c thnh phn cu to v chc nng ging vi cc gen cc loi sinh vt bc cao (c cu to c th phc tp hn). Nhng nguyn nhn chnh ca nhng hoi nghi ny c l l do nhng hiu bit v ADN vo thi im cn rt hn ch.

Bng chng ADN virut lp trnh t bo

Kt qu

Mt bng chng khc cng c cho vic xc nh ADN l vt cht di truyn bt ngun t cc nghin cu cc virut ly nhim vi khun (Hnh 16.3). Nhng virut ny cn c gi l
Chut sng Chut sng Chut cht

Chut cht

u phag a nn phn ui Si ui ADN

Trong mu mu, c t bo chng S c th sinh sn, to nn cc t bo chng S th h con

Kt lun Griffith kt lun rng vi khun R sng c bin i thnh vi khun S gy bnh bng mt cht di truyn khng bit no bt ngun t cc t bo S cht; iu ny dn n hin thng t bo R tr nn c lp v. Ngun F. Griffith, The significance of pneumococcal types,
Journal of Hygiene 27: 113 - 119 (1928). iu g Nu ? Trn c s no th nghim trn y loi tr sao ? kh nng cc t bo chng R c th ch cn n gin dng

T bo vi khun
Hnh 16.3 Virut ly nhim t bo vi khun. Phag T2 v cc phag c quan h khc tn cng t bo vi khun ch v bm vt cht di truyn ca chng qua mng sinh cht (plasma membrane); trong khi , phn u v ui ca phag c gi li bn ngoi b mt t bo vi khun (nh chp qua knh hin vi in t truyn qua t mu - HVTTQm)

lp v ca cc t bo S cht c th chuyn thnh dng vi khun c (gy bnh)?

306

khi kin thc 3

Di truyn hc

bacteriophag (ngha l th n khun hay thc khun th) hoc c gi tt l phag. So vi cc t bo, cc virut c cu to n gin hn nhiu. Mt virut thng ch bao gm ADN (hoc i khi l ARN) c bao bc bi mt lp v protein. c th sinh sn, virut phi ly nhim vo trong mt t bo ri ginh ly b my trao i cht ca t bo. Cc phag v ang c s dng rng ri lm cng c nghin cu di truyn hc phn t. Nm 1952, Alfred Hershey v Martha Chase tin hnh th nghim cho thy ADN l vt cht di truyn ca phag c tn l T2. y l mt trong nhiu loi virut ly nhim Escherichia coli (E. coli), mt loi vi khun thng sng trong rut ng vt c v. Thi k , cc
Hnh 16.4

nh sinh hc bit r l: phag T2, ging vi nhiu phag khc, c thnh phn cu to hu nh ch gm ADN v protein. H cng ng thi bit rng phag T2 c th nhanh chng chuyn t bo E. coli thnh mt nh my sn xut T2 dn n s gii phng nhiu bn sao phag cng s phn r t bo. Bng mt cch no , T2 c th ti lp trnh t bo ch ca n sn sinh cc virut. Nhng, cu hi l: thnh phn no ca virut - protein hay ADN - chu trch nhim cho qu trnh ? Hershey v Chase tr li cu hi ny bng vic thit k mt th nghim cho thy ch mt trong hai thnh phn ca phag T2 xm nhp c vo trong t bo E. coli trong qu trnh ly nhim (Hnh 16.4). Trong th nghim ca mnh, cc

Nghin cu pht hin

Protein hay ADN l vt cht di truyn ca phag T2?

Th nghim Alfred Hershey v Martha Chase s dng cc ng v phng x 35S v 32P nhm tng ng xc nh "s phn" bin i ca cc protein v ADN c ngun gc phag T2 sau khi chng ly nhim vo t bo vi khun. H mun xc nh phn t no trong cc phn t ny i vo t bo v ti lp trnh hot ng ca vi khun gip chng c th sn sinh ra nhiu virut th h con.
Phag nh du phng x c trn vi vi khun. Phag ly nhim cc t bo vi khun. Protein c nh du phng x Khuy mnh hn hp bng my xay lm tung phn phag bn ngoi t bo ra khi t bo. V protein Ly tm cc t bo vi khun dnh kt vi nhau thnh cn ly tm y ng nghim; phn bn ngoi ca phag v phag t do nh hn nn dng phn tn trong dch ly tm. o hot phng x trong phn cn ly tm v dch ly tm. Hot phng x (protein phag) c trong dch ly tm.

Phag T bo vi khun

ADN L th nghim 1: Phag c nui trong mi trng cha ng v phng x lu hunh (35S) nh du protein ca phag (mu hng). ADN c nh du phng x ADN phag

Ly tm Cn ly tm (gm t bo vi khun v cc thnh phn ca n)

L th nghim 2: Phag c nui trong mi trng cha ng v phng x phospho (32P) nh du ADN ca phag (mu xanh dng).

Ly tm Cn ly tm Hot phng x (ADN phag) c trong cn ly tm.

Kt qu Khi protein c nh du (l th nghim 1), hot tnh phng x c gi bn ngoi t bo; nhng khi ADN c nh du phng x (l th nghim 2), hot tnh phng x c tm thy bn trong t bo. Cc t bo vi khun mang ADN ca phag nh du phng x gii phng ra cc virut th h con mang ng v phng x 32P. Kt lun ADN ca phag i vo t bo vi khun, nhng protein ca phag th khng. Hershey v Chase kt lun rng: ADN, ch khng phi protein, c chc nng l vt cht di truyn phag T2. Ngun A.D. Hershey and M. Chase, Independent functions of viral protein and nucleic acid in growth of bacteriophage, Journal of General
Physiology 36: 39 - 56 (1952) iu g Nu ?

Kt qu th nghim s khc bit nh th no nu nh protein l vt cht mang thng tin di truyn?

Chng 16

C s di truyn hc phn t

307

nh khoa hc dng ng v phng x ca lu hunh (S) nh du protein trong mt l th nghim, v s dng ng v phng x ca phospho (P) nh du ADN trong l th nghim th hai. Bi v protein cha lu hunh trong thnh phn cu to ca n, trong khi ADN th khng, nn cc nguyn t S phng x ch kt hp vo cc phn t protein ca phag. Tng t nh vy, cc nguyn t P phng x ch nh du ADN, m khng nh du protein, bi v hu ht cc nguyn t phospho ca phag u trong phn t ADN ca n. Trong th nghim ny, cc nh khoa hc cho cc t bo E. coli khng nh du phng x ly nhim c lp vi phag T2 thu c t hai l th nghim nh du phng x protein v ADN. Cc nh khoa hc sau kim tra xem loi phn t no - ADN hay protein - i vo cc t bo vi khun ngay sau qu trnh ly nhim, qua n c kh nng ti lp trnh hot ng ca cc t bo. Hershey v Chase pht hin ra rng chnh ADN ca phag i vo t bo vi khun, trong khi protein ca phag th khng. Hn na, khi cc t bo vi khun ny c cy chuyn tr li mi trng nui cy, s ly nhim vn tip tc din ra, v cc t bo E. coli gii phng ra cc phag mang mt phn cc nguyn t P phng x. iu ny cho thy thm rng ADN trong t bo gi mt vai tr lin tc trong qu trnh ly nhim. Hershey v Chase kt lun rng ADN c phag tim vo vi khun phi l phn t mang thng tin di truyn t t bo vi khun mi c th to nn cc ADN v protein mi ca virut. Nghin cu ca Hershey v Chase c tnh bc ngot, bi v n cung cp mt bng chng rt thuyt phc rng cc axit nucleic, ch khng phi protein, l vt cht di truyn, t nht l virut.

Khung ng phosphate
u 5'

Cc baz nit

Phosphate ng (deoxyribose) u 3'

Cc bng chng khc chng minh ADN l vt cht di truyn

Cc bng chng khc chng minh ADN l vt cht di truyn n t phng th nghim ca nh ha sinh hc Erwin Chargaff. ADN c bit l polyme ca cc nucleotide. Trong , mi nucleotide gm c 3 thnh phn: mt baz cha nit (gi tt l baz nit; i khi l baz nitric), mt ng pentose c gi l deoxyribose, v mt nhm phosphate (Hnh 16.5). Cc baz c th l adenine (A), thymine (T), guanine (G) hay cytosine (C). Chargaff phn tch thnh phn baz ca ADN t nhiu sinh vt khc nhau. Nm 1950, ng cng b thnh phn baz trong ADN bin ng khi so snh gia cc loi khc nhau. Chng hn, ngi 30,3% cc nucleotide ADN cha baz A, trong khi t l ny E. coli ch l 26%. Bng chng v tnh a dng phn t nh vy gia cc loi, vn trc cha tng bit i vi ADN, cng c thm nhn nh ADN l nhm cht c tim nng hn trong vai tr vt cht di truyn. Chargaff cng c bit nhn mnh mt qui lut k l v t l gia cc baz nit mi loi. Trong thnh phn ADN ca tt c cc loi c nghin cu, s lng adenine lun xp x thymine, cn s lng guanine lun xp x cytosine. Chng hn, trong ADN ca ngi t l ca bn loi baz nit c xc nh l: A = T = 30,3%; G = 19,3% v C = 19,9%. S cn bng v s lng baz A vi T cng nh gia G vi C cn c gi l lut Chargaff. C s phn t ca lut Chargaff trong thc t tn ti nh mt b n cho n khi Watson v Crick pht hin ra cu trc chui xon kp vo nm 1953.

Nucleotide ca ADN

Hnh 16.5 Cu trc mt mch ADN. Mi nucleotide n phn cha mt baz nit (T, A, C hoc G), ng deoxyribose (mu xanh dng) v mt nhm phosphate (mu vng). Nhm phosphate ca nucleotide ny lin kt vi ng ca nucleotide tip theo to nn "ct sng" phn t gm cc nhm phosphate v ng lun phin, t cc baz nh ra. Mch polynucleotide c tnh nh hng t u 5' (vi nhm phosphate) ti u 3' (vi nhm -OH). 5 v 3 l cc s ch cc nguyn t carbon nm trn cu trc vng ca phn ng.

Xy dng m hnh cu trc ADN: Qu trnh tm hiu khoa hc

Sau khi cc nh khoa hc c thuyt phc bi cc bng chng chng minh ADN l vt cht di truyn, mt thch thc c t ra l cn xc nh c cu trc ca ADN t c th gii thch c vai tr di truyn ca n. Vo u nhng nm 1950, s sp xp ca cc lin kt cng ha tr trn mt phn t polyme axit nucleic c bit r (xem Hnh 16.5); do vy, cc nh nghin cu tp trung vo vic lm sng t cu trc khng gian ba chiu ca ADN. Trong cc nh khoa hc nh vy c Linus Pauling t Vin Cng ngh California v Maurice Wilkins cng Rosalind Franklin t i hc King London. Tuy vy, nhng ngi u tin a ra cu tr li ng li l hai nh khoa hc t c bit n vo thi k - James Watson (ngi M) v Francis Crick (ngi Anh).

308

khi kin thc 3

Di truyn hc

S hp tc ngn ngi nhng rt ni ting ca h gip lm sng t cu trc b n ca ADN ngay sau khi Watson c chuyn thm i hc Cambridge l ni m Crick ang nghin cu cc cu trc protein bng mt k thut gi l tinh th hc tia X (xem Hnh 5.25). Khi thm phng th nghim ca Maurice Wilkins, Watson nhn thy hnh nh nhiu x tia X ca ADN do mt ng nghip qu c ca Wilkins l Rosalin Franklin chp c (Hnh 16.6a). Cc bc nh c to ra bng k thut tinh th hc tia X cho thy chng khng phi cc hnh nh ca cc phn t thc s. Cc im chm v vt nhe nh trn Hnh 16.6b c to ra l do tia X b nhiu x (khc x) khi chng i qua cc si ADN tinh sch xp thng hng. Cc nh khoa hc v tinh th hc thng dng cc cng thc ton hc chuyn ti cc thng tin t cc hnh nh nh vy thnh hnh dng ba chiu ca cc phn t; ring Watson th quen thuc vi nhng hnh nh c to ra bi cc phn t dng chui xon. Khi nghin cu k nh nhiu x tia X ca ADN do Franklin chp, Watson khng ch tm ra ADN c dng chui xon m ng cn c lng c chiu rng ca chui xon v khong cch gia hai baz nit lin k dc trc chui xon. Chnh chiu rng ca chui xon ch ra n c to nn t hai mch, khng ging vi cng b ngay trc ca Linus Pauling v mt m hnh phn t gm 3 mch. S pht hin ra hai mch gii thch cho vic thut ng thng c dng hin nay m t ADN l chui xon kp (Hnh 16.7). Watson v Crick bt u xy dng cc m hnh ca chui xon kp sao cho ph hp vi cc s liu o c qua cc hnh
u 5'

(a) Rosalind Franklin

(b) nh nhiu x tia X ca ADN do Franklin chp

Hnh 16.6 Rosalind Franklin v nh nhiu x tia X ca ADN. Franklin, mt nh khoa hc li lc v tinh th hc tia X,
thc hin mt th nghim quan trng, t chp c bc nh gip Watson v Crick lun ra cu trc xon kp ca ADN. Franklin qua i nm 1958 do bnh ung th khi c mi 38 tui. ng nghip ca c l Maurice Wilkins c nhn ng gii thng Nobel nm 1962 cng vi Watson v Crick.

nh nhiu x tia X, t tm ra cu trc ha hc ca ADN. ng thi sau khi c mt bn bo co thng nin cha c cng b v nghin cu ca Franklin, hai nh khoa hc bit rng Franklin tng kt lun rng khung ng - phosphate nm bn ngoi chui xon kp. S sp xp ny rt thuyt phc v n

Lin kt hydro

u 3'

u 3' u 5' (a) Cu trc c bn ca ADN (b) Cu trc ha hc mt on ngn ca ADN (c) M hnh lp kn khng gian Hnh 16.7 Chui xon kp. (a) Di "ruy bng" trong hnh v biu din khung ng - phosphate ca hai mch n ADN. Chui xon ny theo "chiu phi", ngha l vn v pha phi theo hng i ln. Hai mch chui xon c gi li vi nhau qua cc lin kt hydro (ng nt chm mu ) hnh thnh gia cc baz nit kt thnh tng cp bn trong chui xon. (b) thy cu trc ha hc r hn, hai mch ADN c v thng hng nh mt on ngn ca chui xon. Lin kt cng ha tr mnh ni cc tiu n v (nucleotide) trn mt mch vi nhau, trong khi cc lin kt hydro yu hn gi hai mch n vi nhau. iu ng lu l hai mch n ny l i song song, ngha l chy song song nhng theo 2 chiu ngc nhau. (c) S xp chng ln nhau ca cc cp baz nit mt cch cht ch c thy r trong m hnh c v bi my tnh ny. Lc hp dn Van de Waals gia tng cp baz xp chng ln nhau chnh l yu t chnh gip gi ton b phn t vi nhau (xem Chng 2).
Chng 16

C s di truyn hc phn t

309

s y cc baz nit c tnh k nc tng i vo trong v ri xa khi cc phn t nc trong phn dung dch bao quanh. Watson xy dng c mt m hnh vi cc baz nit hng vo pha trong ca chui xon kp. Trong m hnh ny, hai khung ng - phosphate chy i song song vi nhau, ngha l cc tiu n v ca chng chy song song nhng ngc chiu nhau (xem Hnh 16.7). Chng ta c th tng tng s sp xp tng th ging nh mt chic thang dy vi cc thanh thang vng chc. Hai dy ca thang hai bn tng ng vi hai khung ng - phosphate, cn mi thanh thang tng ng vi mt cp baz nit. Lc ny, chng ta c th tng tng mt u ca thang dy c c nh, cn u kia c vn xon, to nn cu trc xon l xo u n. Cc s liu ca Franklin ch ra rng chiu di mi vng xon trn vn dc trc chui xon di 3,4 nm. Vi khong cch gia hai lp cp baz k tip xp chng ln nhau l 0,34nm, s c 10 lp cp baz nit (tng ng vi 10 thanh thang) trong mi vng ca chui xon. Cc baz nit trong chui xon kp kt cp vi nhau theo t hp c hiu: adenine (A) vi thymine (T), cn guanine (G) vi cytosine (C). Th nghim theo m hnh "th v sai", Watson v Crick pht hin c c im quan trng ny ca ADN. u tin, Watson tng tng cc baz kt cp theo tng loi, ngha l A vi A, C vi C. Nhng m hnh ny khng ph hp vi cc s liu tia X vn cho bit ng knh dc chui xon l ng nht. V sao s sp xp ny khng ph hp vi kiu kt cp theo tng loi? Adenine v guanine l cc purine; cc baz ca chng c cu trc hai vng hu c. Ngc li, cytosine v thymine thuc mt h cc baz khc gi l pyrimidine vn ch c cu trc mt vng. Do vy, cc purine (A v G) s c chiu rng gp khong 2 ln so vi cc pyrimidine (C v T). Mt cp purine - purine s l qu rng, trong khi mt cp pyrimidine pyrimidine s l qu hp, so vi ng knh khong 2 nm ca chui xon kp. Nhng, nu s kt cp lun l mt purine vi mt pyrimidine th ng knh chui xon s ng nht.
Purine + Purine: qu rng

ng ng

ng ng

Hnh 16.8 S kt cp cc baz nit trong ADN. Cc cp baz nit trong mt chui xon kp ADN c gi li vi nhau bi cc lin kt hydro (trn hnh y c v bng ng nt chm mu ).

Pyrimidine + Pyrimidine: qu hp

ng. Do vy, trn ADN ca tt c cc sinh vt (ch tr cc virut c vt cht di truyn khng phi ADN si kp), s lng adenine lun bng thymine, cn s lng guanine lun bng cytosine. Mc d nguyn tc kt cp ca cc baz nit l c s to nn cc thanh thang ca chui xon kp, nhng n khng hn ch trnh t ca cc nucleotide dc theo chui xon. Trt t ca bn loi baz nit dc theo chui xon c th bin i bt k, nhng mi gen ch c mt trt t duy nht ca cc chng; trt t ny c gi l trnh t cc baz nit. Thng T nm 1953, Watson v Crick lm sng st cng ng khoa hc bng vic cng b mt bi bo ngn ch di 1 trang trn tp ch Nature*. Bi bo m t m hnh ADN m h tm ra: chui xon kp, ri t n dn tr thnh biu tng ca sinh hc phn t. V p ca m hnh ny chnh l ch: cu trc ADN ch cho chng ta thy c ch sao chp ca n.
Kim tra khi nim

16.1

Purine + Pyrimidine: chiu rng ph hp vi s liu tia X

1. Mt loi rui c t l cc nucleotide trong ADN nh

sau: 27,3% A, 27,6% T, 22,5% G v 22,5% C. Nguyn tc Chargaff c chng minh bi cc s liu trn nh th no?

Watson v Crick t suy lun rng: hn phi c mt kiu kt cp c hiu b sung no c to ra bi cu trc ca cc baz nit. Mi baz nit u c cc gc ha hc mt bn c th to lin kt hydro vi cc "i tc" ca chng: Adenine c th to hai lin kt hydro vi thymine v ch vi thymine; trong khi guanine c th to ba lin kt hydro vi cytosine v ch vi cytosine. Ni cch khc, A kt cp vi T, cn G kt cp vi C (Hnh 16.8). M hnh ca Watson v Crick ng thi gii thch c cho nguyn tc Chargaff. Bt c khi no mt mch ca phn t ADN si kp c A, th mch i din s l T; v G c mt trn mt mch s lun lun kt cp vi C trn mch b sung tng

2. M hnh ca Watson v Crick gip gii thch

nguyn tc Chargaff nh th no?

3.

Nu bin np khng xy ra trong th nghim ca Griffith, th kt qu th nghim s c th khc bit nh th no? Gii thch.
Xem gi tr li Ph lc A.

iu g Nu ?

J.D. Watson and F.H.C.Crick, Molecular structure of nucleic acids: a structure for deoxyribose nucleic acids, Nature 171: 737-738 (1953).

310

khi kin thc 3

Di truyn hc

Khi nim

Nhiu protein phi hp trong sao chp v sa cha ADN

Mi quan h gia cu trc v chc nng bc l r nt chui xon kp. Chnh suy ngh cho rng c s kt cp c hiu gia cc baz nit trong phn t ADN lm le ln tng a Watson v Crick n vi m hnh cu trc chnh xc ca chui xon kp. Cng lc , h tm thy ngha v mt chc nng ca nguyn tc kt cp cc baz. H kt thc bi bo ca mnh bng mt cu pht biu hi hc nh sau: iu m nh chng ti l nguyn tc kt cp c hiu m chng ti coi nh nh ngay lp tc ch ra mt c ch sao chp vt cht di truyn c th tn ti. Trong phn ny, chng ta s cp n nguyn l c bn ca s sao chp (ti bn) ADN cng nh mt s c im chi tit quan trng ca qu trnh .

16.2

Nguyn l c bn: kt cp baz vi mch lm khun

Trong mt bi bo th hai, Watson v Crick pht biu gi thit ca h v c ch sao chp ADN nh sau:
Lc ny, trong thc t m hnh v axit deoxyribonucleic ca chng ti chnh l mt cp cc mch lm khun, m mi mch b sung vi mch cn li. Chng ti tng tng ra rng: trc khi s sao chp din ra, cc lin kt hydro b t g y, sau hai mch gi n xon v tch khi nhau. Mi mch sau c dng lm khun hnh thnh nn mt mch mi i km vi n; nh vy, cui cng chng ta s nhn c hai cp chui xut pht t mt cp chui ban u. Ngoi ra, trnh t ca cc cp chui baz c sao chp mt cch chnh xc.*
*

F.H.C.Crick and J.D. Watson, The complementary structure of deoxyribonuleic acid, Proceedings of the Royal Society of London A 223: 80 (1954).

Hnh 16.9 m t tng c bn ca Watson v Crick. d tng tng, trn hnh ch minh ha mt on chui xon kp ngn dng dui thng. iu ng lu l nu chng ta ch bit trnh t ca mt trong hai mch ADN c v trn Hnh 16.9a, th chng ta c th xc nh c trnh t baz nit ca mch cn li trn c s p dng nguyn tc kt cp b sung ca cc baz. Ni cch khc, hai mch ADN b sung cho nhau; mi mch mang thng tin cn thit ti thit mch cn li. Khi mt t bo sao chp mt phn t ADN, mi mch ca phn t ADN c dng lm khun sp xp cc nucleotide vo mch mi b sung vi n. Cc nucleotide xp hng dc theo mch lm khun tun th nguyn tc kt cp ca cc baz ng thi lin kt vi nhau hnh thnh nn mt mch mi. Nu c mt phn t ADN si kp vo u qu trnh sao chp, th s nhanh chng thu c mt bn sao chnh xc ca phn t m. C ch sao chp ging nh vic dng phim m bn to nn nh dng bn; sau nh dng bn li c dng to nn mt phim m bn mi ri qu trnh c lp i lp li. M hnh sao chp ADN nh vy khng c kim chng trong nhiu nm k t khi cu trc ca ADN c cng b. Th nghim nhm chng minh cho m hnh ny v nguyn tc th d tng tng, nhng v thc nghim th kh thc hin. M hnh ca Watson v Crick d on rng khi chui xon kp sao chp, mi phn t con s mang mt mch c c ngun gc t phn t m cn mt mch c tng hp mi. M hnh bn bo ton ny nh vy khng ging vi m hnh bo ton vn cho rng sau qu trnh sao chp bng mt cch no cc mch ca chui xon kp kt cp tr li vi nhau (tc l phn t ADN m c bo ton nguyn vn). M hnh ny ng thi cng khc vi m hnh phn tn l m hnh cho rng c bn mch ADN sau qu trnh sao chp l s t hp li ca cc phn on ADN c xen ln cc phn on ADN mi tng hp (Hnh 16.10 trang sau). Mc d c ch ADN c th sao chp theo kiu bo ton hoc phn tn l kh gii thch, nhng trong thc t khng th loi b nhng m hnh ny nu thiu bng chng

(a) Phn t ADN m c hai mch ADN b sung vi nhau. Mi loi baz kt cp c hiu vi mt loi baz tng ng ca n qua cc lin kt hydro; trong A lin kt vi T, v G lin kt vi C.

(b) Bc u tin trong sao chp l hai mch n ADN tch nhau ra. Mi mch ca phn t ADN m lc ny c dng lm khun xc nh trnh t cc nucleotide dc theo mch ADN mi, b sung vi n.

(c) Theo nguyn tc b sung, cc nucleotide "xp hng" dc mch mi v lin kt vi nhau to nn khung ng - phosphate. Mi phn t "con" s c mt mch c ca phn t ADN m (xanh sm) v mt mch ADN mi (xanh nht).

Hnh 16.9 M hnh sao chp ADN c bn. Trong m hnh gin lc ny, mt phn on ADN si kp ngn c gin xon thnh dng cu trc ging nh mt chic "thang dy". Trong "dy thang" hai bn l khung ng - phosphate trn hai mch ADN; cn mi "thanh thang" l mt cp baz nit trn hai mch lin kt hydro vi nhau theo nguyn tc b sung (nguyn tc Chargaff). Cc hnh n gin biu din bn loi baz. Trong , mu xanh sm biu din cc mch ADN c ngun gc t phn t ADN m; cn mu xanh nht biu din cc mch ADN c tng hp mi.
Chng 16

C s di truyn hc phn t

311

T bo m (a) M hnh bo ton. Hai mch lm khun kt hp tr li vi nhau sau qu trnh sao chp; v vy, si xon kp "m" c khi phc li nh ban u.

Sao chp ln I

Sao chp ln II

Hnh 16.11

Nghin cu pht hin

ADN sao chp theo kiu bo ton, bn bo ton hay phn tn?
Th nghim Ti Vin cng ngh California, Mathew Meselson v Franklin Stahl nui cy t bo E. coli qua mt s th h trong mi trng cha cc nucleotide tin cht c nh du bng ng v phng x nng 15N. Cc nh khoa hc sau chuyn vi khun sang mi trng ch cha ng v nh 14N. Sau 20 pht v 40 pht, cc mu vi khun nui cy c ht ra tng ng vi hai ln sao chp ADN. Meselson v Stahl c th phn bit c cc phn t ADN c t trng khc nhau bng phng php li tm sn phm ADN c chit rt t vi khun. Nui vi khun trong mi trng cha 15N. Kt qu Li tm mu ADN sau 20 pht (sao chp ln I). Kt lun Li tm mu ADN sau 40 pht (sao chp ln II). T trng thp T trng cao Chuyn vi khun sang mi trng cha 14N.

(b) M hnh bn bo ton. Hai mch ca si xon kp "m" tch nhau ra; mi mch c dng lm khun tng hp nn mt si kp mi .

(c) M hnh phn tn. Mi mch ca hai phn t ADN si kp "con" u l hn hp ca cc phn on c xen ln cc phn on mi tng hp. Hnh 16.10 Ba m hnh sao chp ADN. Mi on xon kp ngn trn hnh biu din mt phn t ADN trong t bo. Bt u t t bo "m" ban u, chng ta theo di qu trnh sao chp hnh thnh nn hai th h t bo "con" tng ng vi hai ln sao chp ADN. Mu xanh nht biu din cc on ADN c tng hp mi.

Meselson v Stahl so snh kt qu thc nghim ca h vi kt qu d on tng ng vi cc m hnh l thuyt (Hnh 16.10) c minh ha di y. Ln sao chp u tin (ln I) to ra mt bng ADN lai "15N-14N" duy nht. Kt qu ny loi b m hnh sao chp kiu bo ton. Ln sao chp th hai (ln II) to ra mt bng ADN nh v mt bng ADN lai. Kt qu ny loi b m hnh sao chp theo kiu phn tn. Trn c s , cc nh khoa hc kt lun rng ADN sao chp theo kiu bn bo ton. Sao chp ln I Sao chp ln II

M hnh bo ton

thc nghim. Phi n cui nhng nm 1950, sau khong 2 nm nghin cu, Matthew Meselson v Franklin Stahl mi thit k c mt m hnh th nghim sng to gip phn bit c ba m hnh sao chp ADN. Th nghim ca h chng minh m hnh sao chp ADN theo kiu bn bo ton c Watson v Crick d on l ng. M hnh th nghim ca Meselson v Stahl sau c cc nh sinh hc coi nh mt v d in hnh v thit k th nghim hp l (Hnh 16.11). Mc d v nguyn tc, c ch sao chp ADN l n gin, nhng trong thc t qu trnh ny din ra lin quan n nhiu s kin phc tp nhng hi ha c cp di y.

M hnh bn bo ton

M hnh phn tn

Ngun M. Meselson and F.W. Stahl, The replication of DNA in

Escherichia coli, Proceedings of the National Academy of Sciences USA 44: 671 - 682 (1958). Thc hnh iu tra c v phn tch bi bo gc trong tiu mc Thc hnh iu tra: hiu v din gii cc bi bo khoa hc. iu g Nu Nu Meselson v Stahl bt u nui vi khun trong mi trng cha 14N ri sau mi chuyn vi khun sang mi trng cha 15N, kt qu s nh th no?

Sao chp ADN: quan st gn hn

Vi khun E. coli c mt nhim sc th duy nht cha khong 4,6 triu cp nucleotide. Trong iu kin mi trng thun li, mi t bo E. coli c th sao chp ton b phn t

312

khi kin thc 3

Di truyn hc

ADN ny v phn chia thnh hai t bo con c vt cht di truyn ging ht nhau trong vng cha n 1 gi. Mi t bo trong c th bn c 46 nhim sc th trong nhn t bo; trong , mi nhim sc th l mt phn t ADN xon kp di, mch thng. Tnh tng cng, h gen nhn ngi cha khong 6 t cp baz, tc l ln hn h gen ca t bo vi khun. Nu chng ta in ton b trnh t h gen ngi di dng cc k t A, T, G v C kch c nh trnh by y, th thng tin di truyn trong 6 t cp nucleotide trong mt t bo lng bi s tng ng vi khong 1200 cun sch c kch thc nh cun sch ny. Vy m, mi t bo ch cn vi gi c th sao chp ton b lng thng tin vi mc sai st rt thp (tn s sai st ch vo khong 10-9, ngha l c 1 t nucleotide mi c mt nucleotide sao chp sai). Ni cch khc, s sao chp ADN l mt qu trnh din ra vi tc cc k nhanh v chnh xc. C hng chc enzym v protein tham gia vo qu trnh sao chp ADN. Nhng hiu bit n nay v b my sao chp vi
im khi u sao chp

khun (chng hn nh E. coli) l y hn so vi sinh vt nhn tht (eukaryote). Nu khng c ch gii g thm, cc bc c m t y l qu trnh xy ra E. coli. Mc d vy, cc nghin cu cho n nay nhn chung cho thy phn ln cc nguyn tc sao chp ADN c bn l ging nhau sinh vt nhn s (prokaryote) v sinh vt nhn tht (eukaryote).

S khi u sao chp

Qu trnh sao chp ADN lun bt u ti nhng v tr c th trn phn t ADN c gi l im khi u sao chp. l nhng on ADN ngn c trnh t nucleotide xc nh. Ging nhiu vi khun ni chung, nhim sc th ca E. coli c dng vng v ch c mt im khi u sao chp. Cc protein khi u sao chp nhn ra v tr ny v gn vo ADN; chng tch hai mch ADN ra khi nhau v to nn bng sao chp. T im khi u sao chp, qu trnh sao chp ADN tin v c hai pha (Hnh 16.12a) cho n khi ton b phn t ADN c
im khi u sao chp Phn t ADN si xon kp Mch lm khun (m) Mch mi tng hp (con)

Mch lm khun (m) Mch mi tng hp (con) Chc sao chp Bng sao chp

Phn t ADN si xon kp

Bng sao chp

Chc sao chp

Hai phn t ADN con Hai phn t ADN con

(a) Nhim sc th dng vng E. coli v nhiu vi khun khc ch c mt im khi u sao chp. Ti im khi u sao chp, hai mch n ADN tch khi nhau hnh thnh nn bng sao chp gm hai chc sao chp. Qu trnh sao chp tin v c hai pha cho n khi cc chc sao chp hai u gp nhau; kt qu to nn hai phn t ADN con. Hnh nh chp bng knh hin vi in t truyn qua (TEM) trn cho thy nhim sc th vi khun ch c mt bng sao chp.

(b) Nhim sc th eukaryote l nhng phn t ADN dng mch thng, kch thc ln. Qu trnh sao chp ADN bt u khi bng sao chp hnh thnh ti nhiu im dc phn t ADN. Bng sao chp m rng khi hai chc sao chp ca n tin dn v hai pha. Cui cng, cc bng sao chp dung hp vi nhau v s tng hp cc mch ADN con kt thc. Hnh nh chp bng TEM trn cho thy on nhim sc th trn ca t bo chut Hamster dng Chinese c ba bng sao chp. Trn nh TEM hnh (b), hy v thm mi tn bng sao chp th ba.
Chng 16 V tip

Hnh 16.12 Cc im khi u sao chp E. coli v sinh vt nhn tht (eukaryote). Mi tn mu ch s dch chuyn ca chc sao chp, ngha l chiu sao chp ADN din ra mi bng sao chp.

C s di truyn hc phn t

313

sao chp xong. Khng ging vi khun, nhim sc th sinh vt nhn tht c th c hng trm, thm ch hng nghn im khi u sao chp. Kt qu l nhiu bng sao chp c hnh thnh; cui cng chng dung hp vi nhau to nn nhng bn sao hon chnh ca cc phn t ADN c kch thc rt ln (Hnh 16.12b). Bng cch ny, tc sao chp ADN sinh vt nhn tht tng ln. Cng ging vi khun, qu trnh sao chp ADN sinh vt nhn tht tin v c hai pha ca bng sao chp. hai u ca bng sao chp c chc sao chp. l vng c dng ch Y l ni hai mch n ADN ca chui xon kp tch nhau ra. C mt s protein tham gia vo hot ng tho xon ny (Hnh 16.13). Helicase l nhm cc enzym c vai tr gin xon v tch hai mch n ra khi nhau. Mi mch n sau c s dng lm khun (mch m) tng hp nn mch ADN mi. Sau khi cc mch lm khun tch nhau ra, cc protein lin kt mch n, gi tt l SSB, s nh kt vo mch ADN lm khun v gip chng tr nn n nh. S tho xon ca chui xon kp trong vng bng sao chp s lm cho cc u chc sao chp tr nn b vn xon cht hn v hnh thnh lc cng. Topoisomerase l nhm enzym gip gii ta lc cng ny bng kh nng lm t gy tm thi cc lin kt cng ha tr trn mch n ADN, xoay cc mch n quanh nhau tho xon, ri ni chng tr li vi nhau. Cc mch ADN m sau khi gin xon c dng lm khun tng hp cc mch ADN mi. Tuy vy, enzym trc tip tng hp ADN khng c kh nng khi u qu trnh tng hp chui polynuclotide mi; chng ch c th b sung cc nucleotide vo u 3 ca mt chui c sn nu nucleotide b sung kt cp ph hp vi nucleotide trn mch ADN lm khun. V l do ny, chui nucleotide u tin c to ra trong tng hp ADN lun l mt on ngn ARN, ch khng phi ADN. on ARN ngn ny c gi l on mi v c xc tc tng hp bi enzym primase (xem Hnh 16.13). Primase bt u tng hp on mi t mt ribonucleotide (hay nucleotide ARN) duy nht; sau , mi ln phn ng n gn thm mt ribonucleotide theo nguyn tc b sung vi mch ADN lm khun. Mt on mi hon chnh, gm khong 5 - 10 nucleotide, lc ny s bt cp vi mch khun. Mch ADN c tng hp mi s bt u t u 3 ca on mi ARN.
Hnh 16.13 Mt s protein lin quan n khi u sao chp ADN. Cc loi protein ging nhau hot ng c hai chc sao chp ca cng mt bng sao chp. gin lc, y ch minh ha mt chc sao chp.

Tng hp mch ADN mi

Cc enzym c tn gi l ADN polymerase chnh l cc enzym trc tip xc tc tng hp mch ADN mi bng vic b sung cc nucleotide vo mt chui sn c. E. coli, c mt s loi ADN polymerase khc nhau; tuy vy, hai loi c vai tr chnh trong sao chp ADN l ADN polymerase III v ADN polymerase I. c im ny sinh vt nhn tht l phc tp hn. n nay, c t nht 11 loi ADN polymerase khc nhau c xc nh; tuy vy, nguyn l hot ng chung ca chng v c bn l ging nhau. Phn ln cc enzym ADN polymerase u cn mt on mi v mt mch ADN lm khun. Trn c s trnh t ca mch lm khun, chng b sung cc nucleotide mi dc theo mch c tng hp mi theo nguyn tc b sung. E. coli, ADN polymerase III (vit tt l ADN pol III) b sung cc nucleotide ADN vo on mi ri tip tc ko di chui ADN theo nguyn tc b sung vi mch lm khun cho n khi kt thc chui. Tc ko di chui ADN vo khong 500 nucleotide mi giy vi khun, v vo khong 50 nucleotide mi giy ngi. Mi nucleotide khi c b sung vo chui ADN ang ko di u dng nucleotide triphosphate; l mt nucleoside (gm ng pentose v baz nit) lin kt vi ba nhm phosphate. Chng ta cp n mt phn t nh vy l ATP (adenosine triphosphate; xem Hnh 8.8). S khc bit duy nht gia phn t ATP (c vai tr trong trao i nng lng) vi dATP (l tin cht ca adenine trong ADN) l thnh phn ng. Nu nh ng trong ADN l deoxyribose th ng trong ATP l ribose. Cng ging nh ATP, cc nucleoside triphosphate c dng tng hp nn ADN l nhng cht ha hc phn ng mnh; mt phn bi chng cha ui triphosphate vn tch in m v km bn. Mi ln mt nucleotide gn thm vo chui ang ko di, hai nhm phosphate (k hiu l -i v cn c gi l pyrophosphate) s t khi phn t nucleoside triphosphate tin cht. S thy phn din ra ngay sau ca nhm pyrophosphate thnh hai phn t phosphate v c i i lin vi cc phn ng sinh nhit l ng lc phn ng trng hp ADN din ra (Hnh 16.14).
Primase tng hp on mi ARN c trnh t b sung vi mch khun.

Cc protein lin kt mch n (SSB) gip lm n nh mch ADN lm khun.

Topoisomerase lm t gy cc mch n, tho xon ri ni chng tr li vi nhau. on mi ARN

Helicase lm gin xon v tch hai mch n ADN khi nhau.

314

khi kin thc 3

Di truyn hc

Mch mi u 5'

Mch khun u 3'

u 5'

u 3'

ng Phosphate

Hnh 16.14 S kt hp nucleotide vo mch ADN. Enzym ADN polymerase xc tc vic b sung mt nucleoside triphosphate vo u 3 ca mt mch ADN ang ko di, vi s gii phng hai nhm phosphate.

S dng s ny gii thch ti sao chng ta ni mi mch ADN c tnh phn cc.

u 3'

ADN polymerase

Pyrophosphate u 3' Nucleoside triphosphate

u 5'

u 5' Bng sao chp ADN tng qut im khi u sao chp Mch dn u Mch ra chm Mi Mch ra chm Mch dn u Chiu sao chp chung im khi u sao chp

Ko di chui kiu i song song

Nh nu trn, hai u ca mt mch ADN l khc nhau, to cho mi mch ADN c tnh phn cc, ging nh ng mt chiu vy (xem Hnh 16.5). Ngoi ra, hai mch ADN trong chui xon kp l i song song, ngha l chng phn cc theo chiu i din nhau, cng ging nh hai ln ng mt chiu trn xa l theo hng ngc nhau vy (xem Hnh 16.14). R rng l hai mch mi c tng hp trong qu trnh sao chp ADN phi i song song so vi cc mch khun ca chng. S sp xp i song song ca chui xon kp nh hng th no n qu trnh sao chp? Do c im cu trc, cc enzym ADN polymerase ch c th b sung cc nucleotide vo pha u 3 t do ca mt on mi hoc ca mt mch ADN ang ko di, ch khng bao gi b sung c cc nucleotide vo pha u 5 (xem Hnh 16.14). V vy, mt mch ADN mi ch c th ko di theo chiu 5 3. Vi nguyn tc , hay xem s sao chp din ra th no ti mt chc sao chp (Hnh 16.15). Dc theo mt mch khun ADN, ADN polymerase III c th tng hp mch mi mt cch lin tc theo nguyn tc b sung bng vic ko di mch mi theo chiu bt buc 5 3. ADN pol III mt cch n gin lch vo chc sao chp trn mch khun ri b sung lin tc cc nucleotide vo mch mi cng vi vic chc sao chp tin v pha trc. Mch ADN mi c tng hp theo kiu ny c gi l mch dn u. tng hp mch dn u, ADN pol III ch cn mt on mi duy nht (xem Hnh 16.15). c th ko di mch ADN mi cn li theo ng chiu 5 3, ADN pol III phi hot ng dc theo mch khun cn li theo chiu ngc hng vi chiu dch chuyn ca chc sao chp. Mch ADN mi c tng hp theo chiu ngc hng ny c gi l mch ra chm hay mch theo sau *. Khng ging mch dn u c tng hp lin tc, mch ra chm c tng hp gin on thnh cc on nh. Cc on ca
*

Sau khi on mi ARN c to ra, ADN pol III bt u tng hp mch dn u

Mi ARN Protein kp trt ADN m ADN pol III

Mch dn u c ko di lin tc theo chiu 5 3 cng vi chiu di chuyn ca chc sao chp Hnh 16.15 Tng hp mch dn u trong sao chp ADN. S ny tp trung vo chc sao chp bn tri ca mt bng sao chp. ADN polymerase III (ADN pol III), c v ging nh bn tay khum hnh chn, nh kt cht ch vi mt protein c gi l kp trt, c v ging nh mt chit bnh vng. Protein kp trt y ADN pol III trt dc mch ADN lm khun.
Chng 16

Qu trnh tng hp mch dn u v mch theo sau din ra ng thi vi tc tng ng. S d gi l mch ra (hay chm mch theo sau) l do s tng hp mch ny din ra chm hn cht t so vi mch dn u; mi phn on mi ca mch ra chm ch c khi u tng hp khi mt on mch khun ADN ti chc sao chp bc l di.

C s di truyn hc phn t

315

Bng sao chp ADN tng qut im khi u sao chp Mch ra chm Mch dn u Mch ra chm Mch dn u

Chiu sao chp chung Primase ni nucleotide ARN vo on mi

Mch khun

ADN pol III b sung cc nucleotide ADN vo mi, hnh thnh on Okazaki 1

mch ra chm nh vy c gi l cc on Okazaki theo tn nh khoa hc Nht bn pht hin ra chng. E. coli cc on Okazaki di khong 1000 n 2000 nucleotide, trong khi sinh vt nhn tht chng di khong 100 n 200 nucleotide. Hnh 16.16 minh ha cc bc ca qu trnh tng hp mch ra chm. Nu nh tng hp mch dn u ch cn mt on mi duy nht, th mi on Okazaki trn mch ra chm u cn ring mt on mi. Mt loi ADN polymerase khc, gi l ADN polymerase I (ADN pol I) s thay th cc nucleotide ARN ca on mi bng cc nucleotide ADN tng ng bng vic b sung tng nucleotide vo u 3 ca on Okazaki lin k (on s 2 trn Hnh 16.16). Tuy vy, ADN pol I khng th ni nucleotide cui cng thuc on ADN va c thay th vi nucleotide u tin ca on Okazaki lin k (on s 1 trn Hnh 16.16). Lc ny, mt enzym khc, gi l ADN ligase, s thc hin nhim v ny; n xc tc phn ng ni khung ng - phosphate ca tt c cc on Okazaki vi nhau to nn mch ADN ra chm lin tc. Hnh 16.16 v Bng 16.1 m t tm tt qu trnh sao chp ADN. Hy c v quan st trc khi tip tc cc phn di y.

Phc h sao chp ADN

Khi tip cn on ARN mi k tip, ADN pol III ri khi chc sao chp

on Okazaki

ADN pol III b sung nucleotide vo on mi 2 cho n khi n tip cn on mi 1, ri ri ra.

ADN pol I thay th ARN bng ADN, bng cch ko di u 3 ca on 2.

ADN ligase ni hai on Okazaki vi nhau

y, mch ra chm c sao chp hon chnh

Trc y, d tng tng, cc phn t ADN polymerase c m t ging nh cc u xe la di chuyn dc ng ray ADN; tuy vy, hnh nh khng chnh xc hai im quan trng. Th nht, b my sao chp ADN trong thc t l mt phc h ln gm nhiu protein khc nhau. Cc tng tc protein - protein qui nh hiu qu v chc nng ca phc h ny. Chng hn nh, bng s tng tc vi cc protein khc ti chc sao chp, primase r rng hot ng ging nh mt chic phanh phn t, lm chm s m rng chc sao chp v iu phi tc sao chp tng ng gia mch dn u v mch ra chm. Th hai, phc h sao chp ADN khng di chuyn dc ADN; thay vo , chui ADN c tut qua phc h trong qu trnh sao chp. Trong cc t bo sinh vt nhn tht, nhiu phc h sao chp c l kt nhm vi nhau hnh thnh nn cc nh my; dng cu trc ny, chng c c nh vo mng li nhn (mng li ny c hnh thnh t cc si dn rng qua phn cht nhn ca t bo). Cc nghin cu gn y ng h cho m hnh trong hai phn t ADN polymerase lin kt vo hai mch ADN lm khun (mi phn t enzym lin kt trn mt mch); ri mch ADN lm khun c ko qua enzym ging nh xe ch, kt qu l hai phn t ADN con c hnh thnh v c y ra ngoi. Mt s bng chng b sung cho thy mch ra chm hnh thnh nn cu trc ging thng lng quanh phc h sao chp; v vy, khi ADN polymerase hon thnh vic sao chp mt on Okazaki v ri ra th n khng cch xa ng k so vi on mi ca on Okazaki k tip. Cu trc thng lng ca mch ra chm cho php t bo c th tng hp nhiu on Okazaki trong thi gian ngn.

c sa v sa cha ADN
Chiu sao chp chung Hnh 16.16 Tng hp mch ra chm.

S chnh xc trong sao chp ADN khng n thun ph thuc vo tnh c hiu trong nguyn tc kt cp ca cc baz. Mc d li sao chp ADN xut hin vi tn xut chung vo khong mt trong 10 t nucleotit (10-10), nhng trong thc t cc li kt cp nucleotit ban u vo mch ADN ang m rng bi hot ng ca enzym ADN polymerase thng cao hn khong

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khi kin thc 3

Di truyn hc

Protein lin kt mch n gip mch khun sau khi tho xon tr nn n nh Helicase tho xon si xon kp "m"

Mch dn u c tng hp lin tc theo chiu 5' 3' bi ADN polymerasse

Bng sao chp ADN tng qut im khi u sao chp Mch dn u Mch ra chm Mi Mch dn u Chiu sao chp chung

Mch ra chm

Mch dn u ADN pol III Mi Phn t ADN "m" Primase ADN pol III Mch ra chm ADN pol I ADN ligase

Primase bt u tng hp mi ARN cho on Okazaki th 5 ADN pol III ang hon thnh tng hp on Okazaki th 4. Khi n tip cn mi ca on Okazaki th 3, n s tch ra, ri di chuyn n chc sao chp v tip tc chc nng b sung nucleotit vo u 3' ca on mi thuc on Okazaki th 5. ADN pol I loi b mi u 5' ca on Okazaki th 2, thay th n bng cc nucleotit ADN bng vic b sung tng nucleotit vo u 3' ca on Okazaki th 3. S thay th nucleotit ARN cui cng bng nucleotit ADN s li mt khung ng - phosphate c u 3' t do.
Bng 16.1

ADN ligase ni u 3' ca on Okazaki th 2 vi u 5' ca on Okazaki th nht.

Hnh 16.17 Tm tt qu trnh sao chp ADN vi khun. S ny minh ha mt chc sao chp; nhng nh minh ha trn s tng qut (pha trn bn phi), qu trnh sao chp thng din ra ng thi c hai chc ca mi "bng" sao chp. Ti mi chc sao chp, chng ta d dng nhn thy, mt mch ADN mi c tng hp lin tc v c gi l mch dn u; trong khi mch cn li c tng hp thnh tng on ngn v c gi l mch ra chm.

Protein Helicase

Cc protein sao chp ADN vi khun v chc nng ca chng Chc nng Tho xon chui xon kp ti v tr chc sao chp Lin kt v lm n nh cc mch n ADN cho n khi cc mch ny c dng lm khun cho qu trnh sao chp Lm gim lc cng pha trc chc sao chp bng cch lm t tm thi cc mch ADN, lun chng qua nhau, ri ni li. Tng hp on mi ARN ti u 5 ca mch dn u v ti mi on Okazaki ca mch ra chm S dng mch ADN "m" lm khun, tng hp mch ADN mi bng vic b sung cc nucleotit vo u 3 ca mch ADN sn c hoc on mi ARN qua lin kt cng ha tr Loi b cc nucleotit ARN thuc on mi bt u t u 5, ri thay th chng bng cc nucleotit ARN Ni u 3 ca on ADN c loi b on mi vi phn cn li ca mch dn u, hoc ni gia cc on Okazaki ca mch ra chm C s di truyn hc phn t

Protein lin kt mch n Topoisomerase

100.000 ln - tc l, khong mt nucleotit sai trong c 100.000 nucleotit ca mch lm khun. Trong qu trnh sao chp, cc enzym ADN polymerase c sa tng nucleotit da trn trnh t mch lm khun ngay khi chng b sung thm nucleotit mi vo chui ang ko di. Nu tm ra nucleotit kt cp sai, enzym polymerase s ct b nucleotit ny ri tng hp li bng nucleotit kt cp ng. (Hot ng ny ging nh khi chng ta dng phm "BackSpace" trn bn phm my tnh xa mt k t sai, ri nhp li mt k t ng). Tuy vy, i khi cc nucleotit kt cp sai c th thot khi hot ng c sa ca cc ADN polymerase. Trong c ch sa cha kt cp sai, cc enzym s tin hnh loi b v thay th cc nucleotit sai do cc li ca qu trnh sao chp. Cc nh khoa hc n tm quan trng ca nhng enzym ny khi tm thy mt sai hng di truyn mt trong nhng gen m ha cc enzym nh vy lin quan trc tip n s pht sinh mt dng ung th rut kt. R rng l, sai hng di truyn ny cho

Primase

ADN pol III

ADN pol I

ADN ligase

Chng 16

317

php cc li dn n pht sinh ung th c th tch ly trn phn t ADN vi tc nhanh hn so vi bnh thng. Cc nucleotit kt cp sai hoc bin i cng c th xut hin sau qu trnh sao chp. Trong thc t, duy tr chnh xc thng tin di truyn, cc t bo cn thng xuyn sa cha cc sai hng khc nhau xy ra vi ADN. Cc phn t ADN dng nh lun trng thi bc l vi nhiu nhn t vt l v ha hc nguy hi (s c chng ta cp k hn Chng 17). Cc hp cht phn ng mnh (c mt trong mi trng sng hoc xut hin t nhin trong cc t bo), cc tia phng x, nh sng cc tm v mt s phn t nht nh trong khi thuc l c th gy nn s bin i ca cc nucleotit v nh hng n thng tin di truyn c m ha trn cc phn t ADN. Ngoi ra, bn thn cc baz trong ADN cng c th bin i t pht trong iu kin sinh l bnh thng ca t bo. Tuy nhin, thng th nhng bin i ny s c sa cha cho ng trc khi chng c th tr thnh cc t bin di truyn n nh qua cc th h. Mi t bo u lin tc theo di v sa cha vt cht di truyn ca chng. Do hot ng sa cha ADN c tm quan trng sng cn i vi c th, nn khng c g l ngc nhin khi c nhiu enzym sa cha ADN xut hin trong qu trnh tin ha. E. coli, c khong 100 enzym sa cha ADN c bit n; trong khi , con s ny ngi l khong 130. Phn ln cc h thng sa cha ADN ca t bo, bt k i vi cc sai hng do li sao chp hay cc sai hng khc v cu trc ADN, u da trn nguyn tc b sung gia cc baz trn hai mch ca phn t ADN. Thng thng, mt on trn mch ADN mang nucleotit sai hng c ct b bi mt enzym ct ADN - nuclease - ri on trng hnh thnh s c lp y tr li bng cc nucleotit kt cp ng da trn mch ADN khng b sai hng lm khun. Cc enzym lin quan n vic lp y on trng gm ADN polymerase v ADN ligase. Mt h thng sa cha ADN nh vy c gi l sa cha bng ct b nucleotit (Hnh 16.18). Mt chc nng quan trng ca cc enzym sa cha ADN trong t bo da ca chng ta l sa cha cc sai hng di truyn gy ra do tia cc tm n t nh sng mt tri. Mt loi sai hng nh vy c minh ha trn Hnh 16.18; trong , cc baz thymine lin k vi nhau trn mch ADN hnh thnh lin kt cng ha tr vi nhau. Lin kt kp thymine nh vylm bin dng cu trc ADN bnh thng v nh hng n qu trnh sao chp. Tm quan trng ca sa cha ADN i vi nhng sai hng ny c nhn thy qua bnh kh b sc t; y l bnh gy ra bi sai hng di truyn lin quan n gen m ha enzym sa cha ADN theo c ch ct b nucleotit. Nhng c th ri lon v enzym ny thng rt mn cm vi nh sng mt tri; cc t bin trong t bao da ca h do tia cc tm gy ra vn khng c sa cha, tch ly qua thi gian v c nguy c gy nn bnh ung th da.

Phc kp thymine lm bin dng ADN

Mt enzym nuclease ct on mang nucleotit sai hng trn mt mch ADN ti hai im, ri loi b on sai hng Nuclease

ADN polymerase

ADN polymerase tng hp b cc nucleotit vo on trng, s dng mch khng b sai hng lm khun

ADN ligase

ADN ligase hon thnh vic sa cha ADN bng vic lp lin kt phosphodiester cui cng trn khung ng phosphate

Hnh 16.18 Sa cha ADN kiu ct b nucleotit. Mt nhm cc enzym v protein c vai tr pht hin v sa cha ADN sai hng. Hnh trn cho thy ADN mang mt phc kp thymine, y l mt kiu sai hng ADN thng b gy ra do chiu x UV. Mt enzym nuclease ct b vng ADN sai hng, ri mt enzym ADN polymerase (ADN pol I vi khun) s thay th on b ct bng cc nucleotit ph hp trn c s dng mch ADN khng b sai hng lm khun. Enzym ADN ligase s hon thnh qu trnh sa cha bng cch lp khe h (lin kt phosphodiester) cui cng trn khung ng - phosphate.

Sao chp u tn cng ca phn t ADN

Mc d kh nng sao chp ca cc enzym ADN polymerase l "n tng", nhng trong t bo lun c mt t l nh trnh t ADN m cc ADN polymerase khng th sao chp v sa cha c. i vi cc phn t ADN mch thng, chng hn nh nhim sc th sinh vt nhn tht, cc ADN polymerase ch c th b sung cc nucleotit vo u 3 ca mt chui polynucleotit ang ko di; iu ny dn n vn l: b my sao chp khng c cch no c th sao chp hon chnh phn u 5 ca cc mch ADN con. Ngay c mt on Okazaki c th bt u bng mt on mi ARN lin kt vi u tn cng ca mch ADN lm khun cng khng th thay

th bng ADN bi khng c sn u 3 pha trc phn ng b sung cc nucleotit c th din ra (Hnh 16.19). Kt qu l sau mi ln sao chp, phn t ADN si kp ngy cng ngn li v c cc u khng bng nhau (cn gi l u "ch chi"). Hin tng phn t ADN c xu hng ngn li sau mi ln sao chp thng khng xy ra cc sinh vt nhn s, bi v cc phn t ADN ca chng c dng vng (tc l khng c cc u tn cng). Vy, c ch no bo v cc gen ca sinh vt nhn tht khng mt i sau cc chu k sao chp ADN ni tip nhau ? l do cc phn t ADN nhim sc th sinh vt nhn tht c cc trnh t nucleotit c bit ti cc u tn cng ca chng v c gi l u mt nhim sc th (Hnh 16.20). Vng u mt nhim sc th khng cha cc gen; thay vo , n thng cha cc trnh t nucleotit ngn lp li nhiu ln. Ti cc u mt nhim sc th ngi, mt trnh t ngn gm 6 nucleotit l TTAGGG thng lp li t 100 n 1000 ln. Trnh t ADN ti u mt bo v cc gen ca c th. Ngoi ra, cc protein c

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u mt ca cc mch ADN m

Mch dn u Mch ra chm

on cui Mch dn u Mi ARN

on pha trc

Mch khun Mi c loi b nhng khng c thay th bng ADN do ADN pol khng th hot ng khi thiu u 3 t do

on mi ARN c loi b v thay th bng ADN nh c u 3 ca on pha sau

Hnh 16.20 u mt nhim sc th. Cc sinh vt nhn tht c cc trnh t lp li, khng m ha phn tn cng ca cc phn t ADN mch thng v c gi l u mt. Hnh trn l nhim sc th chut c phn u mt nhum mu vng.

Chu k sao chp th hai

Mch dn u mi Mch ra chm mi Cc chu k sao chp tip theo Cc phn t con ngy cng ngn Hnh 16.19 u mt cc phn t ADN mch thng ngn li sau mi chu k sao chp. Hnh trn ch minh ha mt u ca mt mch phn t ADN si kp qua hai chu k sao chp. Sau chu k th nht, mch ra chm mi ngn hn so vi mch lm khun. Sau chu k th hai, c hai mch dn u v mch ra chm u ngn hn so vi mch ADN m ban u. Mc d khng c v y, nhng hin tng ngn dn cng xy ra vi u mt th hai cn li ca phn t ADN.

hiu lin kt vi ADN ti u mt c vai tr ngn cn cc u "ch chi" ca phn t ADN con khng hot ha cc h thng theo di cc sai hng ADN ca t bo. (Cc u "ch chi" ca phn t ADN, nu hnh thnh do s t gy si xon kp, thng l tn hiu thc y s dng li ca chu k t bo hoc dn n con ng cht theo chng trnh ca t bo). Cu trc u mt nhim sc th khng th gip phn t ADN mch thng trnh khi vic ngn li sao mi chu k sao chp, m chng ch lm chm s "n mn" cc gen gn u tn cng ca cc phn t ADN. Nh minh ha trn Hnh 16.19, u mt nhim sc th ngn li sau mi chu k sao chp. Kt qu l, ADN c xu hng ngy cng ngn hn trong cc t bo soma ang phn chia ngi gi hoc trong cc t bo nui cy tri qua nhiu ln phn bo. Ngi ta cho rng s ngn dn ca u mt cc nhim sc th bng cch no c lin quan trc tip vi qu trnh gi ha nhng m nht nh, thm tr vi s gi ha ca ton b c th. Nhng iu g xy ra vi cc t bo m h gen ca chng vn cn c duy tr nguyn vn qua nhiu th h sinh sn? Nu cc nhim sc th cc t bo mm sinh dc (cc t bo

sinh giao t) tr nn ngn hn sau mi chu k t bo, th nhng gen thit yu cui cng s mt i trong cc t bo giao t m chng sinh ra. Tuy vy, trong thc t, iu ny khng xy ra. C mt enzym, c gi l telomerase, xc tc vic ko di u mt trong cc t bo mm sinh dc sinh vt nhn tht; qua , b p cc nucleotit b mt sau mi chu k sao chp v phc hi li chiu di ban u ca cc phn t ADN. Enzym telomerase khng hot ng hu ht cc t bo soma ngi, nhng hot tnh ca n trong cc t bo mm sinh dc gip u mt cc nhim sc th hp t thng t c di ti a. Vic u mt nhim sc th cc t bo soma thng ngn i sau mi ln phn bo cng c th gip bo v c th khi s pht sinh ung th, bi v qua c ch ny s ln phn bo ca cc t bo soma b hn ch. Cc t bo ca cc khi u ln thng c cc u mt nhim sc th ngn bt thng, c th do chng tri qua nhiu ln phn bo. Nu u mt nhim sc th tip tc ngn i th cc t bo khi u c th cht t pht. Nhng iu ngc nhin l cc nh khoa hc tm thy enzym telomerase hot ng mnh nhiu t bo ung th; iu ny cho thy kh nng ca enzym ny trong vic gip duy tr s n nh u mt nhim sc th cc t bo ung th. Nhiu t bo ung th dng nh c kh nng phn bo khng hn ch, chng hn nh cc dng t bo "bt t" khi c a vo nui cy invitro (xem Chng 12). Nu enzym telomerase l mt nhn t quan trng trong pht sinh nhiu bnh ung th khc nhau, th enzym ny c th l mt "mc tiu" hiu qu trong chn on v iu tr cc bnh ung th tng ng. Ti y, chng ta cp v cu trc v s sao chp ADN si kp. Trong phn tip theo, chng ta s xem ADN c ng gi nh th no trong cc nhim sc th, cu trc ca t bo vn c coi c vai tr mang thng tin di truyn.
Kim tra khi nim

16.2

1. Nguyn tc kt cp b sung ca cc baz nit c vai

tr th no trong sao chp ADN ? 2. Nu hai chc nng chnh ca ADN pol III trong sao chp ADN? Nu ADN pol I b mt chc nng, 3. iu g nu th s sao chp mch dn u s b nh hng nh th no? Trong Bng sao chp tng qut Hnh 16.17, xc nh v tr hot ng ca ADN pol I trn mch dn u .
Xem gi tr li Ph lc A.
Chng 16

C s di truyn hc phn t

319

Khi nim

Mi nhim sc th gm mt phn t ADN c ng gi vi cc phn t protein

Thnh phn chnh trong h gen hu ht vi khun l mt phn t ADN si kp, mch trn lin kt vi mt lng nh protein. Mc d chng ta thng coi cu trc ny l nhim sc th vi khun, nhng thc t cu trc ny rt khc so vi mt nhim

16.3

sc th in hnh sinh vt nhn tht vn thng bao gm mt phn t ADN si kp mch thng lin kt vi mt lng ln protein. E. coli, ADN nhim sc th bao gm khong 4,6 triu cp nucleotit, tng ng vi khong 4400 gen. Lng ADN ny gp khong 100 ln so vi mt h gen virut in hnh, nhng ch bng khong mt phn nghn so vi lng ADN c trong mt t bo soma ngi. D vy, lng ADN vi khun cng l rt ln so vi kch thc ca t bo. Nu dui thng, phn t ADN trong mt t bo E. coli c th o bng n v milimet v di hn khong 500 ln so vi kch thc ca t bo. Tuy vy, trong t bo nh tng tc vi

Hnh 16.21

Khm ph ng gi cht nhim sc trong nhim sc th sinh vt nhn tht

Chui s v nh hin vi in t truyn qua di y m t m hnh biu din cc cp gp xon v siu xon ca nhim sc th. Cc hnh minh ha c phng i t cp cu trc ca mt phn t ADN n l ti nhim sc th k gia nguyn phn l lc nhim sc th co xon cc i v c th quan st c di knh hin vi quang hc thng thng.

Nucleosome (ng knh 10nm) Chui xon kp ADN (ng knh 2nm)

H1 ui histone Cc protein histone

1 Chui xon kp ADN 2 Cc histone

y minh ha m hnh di ruy bng ADN; trong , mi di ruy bng biu din mt khung ng - phosphate. T Hnh 16.7, chng ta nh rng gc phosphate phn b dc khung phn t ny v lm cho phn t ADN c c tnh tch in m sut dc chiu di phn t. nh hin vi in t truyn qua (TEM) trn cho thy mt phn t ADN trn (khng lin kt protein); thit din chiu ngang (ng knh) ca ring chui xon kp ny l 2nm. Cc protein histone c vai tr ng gi ADN vo cht nhim sc cp u tin. Tuy mi phn t histone ch c kch thc nh (khong 100 axit amin), nhng tng khi lng cc histone trong cht nhim sc gn tng ng vi lng ADN. Cc axit amin tch in dng (lysine hoc arginine) chim hn 1/5 tng s cc axit amin c trong histone. Bn loi histone ph bin nht trong cht nhim sc l H2A, H2B, H3 v H4. Cc histone ny rt ging nhau mi sinh vt nhn tht. V d, histone H4 b ch khc histone H4 cy u ng 2 axit amin, cn li l ging ht nhau. S bo th ca protein histone trong sut qu trnh tin ha cho thy vai tr quan trng sng cn ca protein ny trong t chc ADN ca cc t bo sng. Bn loi histone chnh c vai tr quyt nh cp ng gi tip theo ca ADN. (Mt loi histone th 5 l H1 lin quan n mt bc ng gi tip theo ca cht nhim sc).

3 Nucleosome, hay chui

ht (si 10-nm)
Trn nh hin vi in t, si nhim sc cp t chc ny nu khng gp xon c ng knh 10nm (si 10nm). Si nhim sc c dng ging nh mt chui ht vi cc ht xp cch nhau tng i u n. Mi ht l mt nucleosome; y chnh l n v ng gi ADN c bn; si ni gia cc ht c gi l cc on ADN ni. Mt nucleosome lun gm ADN cun quanh li protein 1,65 vng; li protein c cu to t 8 phn t ca 4 loi histone chnh (mi loi ng gp 2 phn t). u amino (u N) tn cng ca mi histone (ui histone) thng th ra ngoi nucleosome. Trong chu k t bo, khi ADN sao chp, cc histone ri khi ADN trong thi gian ngn. Nhn chung, hin tng tng t xy ra khi gen c phin m v b my ca t bo phi tip cn c ADN. Chng 18 s cp n nhng pht hin gn y v vai tr ca nucleosome v ui histone trong iu ha biu hin gen sinh vt nhn tht.

320

khi kin thc 3

Di truyn hc

mt s loi protein nht nh, nhim sc th thng dng gp xon thm ch "siu xon" c th ng gi cht v ch chim mt khong khng gian hn ch trong t bo. Khng ging nhn t bo sinh vt nhn tht, vng cha mt cao ca ADN vi khun khng c lp mng bao bc v ch c gi l vng nhn (xem Hnh 6.6). Mi nhim sc th sinh vt nhn tht u cha mt chui xon kp ADN mch thng duy nht; ngi, kch thc trung bnh vo khong 1,5 x 108 cp nucleotit. Lng ADN nh vy l ln hn nhiu so vi chiu di nhim sc th khi co xon cc i. Nu dui thng, mi phn t ADN h gen nhn ngi c

chiu di trung bnh trn 4 cm, tc l di hn hng nghn ln so vi ng knh ca nhn t bo - l cha k n 45 nhim sc th khc cn li ng thi c mt trong nhn t bo! Trong t bo, ADN ca sinh vt nhn tht kt hp chnh xc vi mt lng ln cc protein. S t hp ca ADN vi cc protein cu trc nhim sc th c gi l cht nhim sc. Nhn t bo c th cha va cht nhim sc l do s ng gi ADN nhiu cp khc nhau bi cc protein cu trc nhim sc th. Hiu bit hin nay ca chng ta v cc cp ng gi ADN c minh ha trn Hnh 16.18. Hy quan st k hnh ny trc khi c cc phn tip theo.

Nhim sc t (700 nm)

Si 30-nm

Si thng lng Khung nhim sc th

Si 300-nm

Nhim sc th nhn i (1400 nm)

4 Si 30-nm
Cp ng gi ADN tip theo l do tng tc gia cc ui histone ca mt nucleosome vi phn ADN ni v vi cc nucleosome lin k hai bn. cp ng gi ny, c s tham gia ca mt histone th nm l H1. Cc mi tng tc ny lm si nhim sc 10nm tip tc cun gp v to nn si c chiu dy khong 30nm (si 30nm). Mc d si 30nm rt ph bin trong k trung gian ca chu k t bo, nhng cn nhiu quan im khc nhau v s sp xp cc nucleosome bc cu trc ny.

5 Min thng lng

(si 300-nm)
Cc si 30nm tip tc cun vng hnh thnh nn dng cu trc ging thng lng, gi l min thng lng,nh vo khung nhim sc th c cu to nn t cc protein. Cu trc ny gi l si 300nm. Khung nhim sc th thng c thnh phn giu v mt loi topoisomerase, ng thi c mt histone H1.

6 Nhim sc th k gia
Trong mt nhim sc th ang nguyn phn, cc min thng lng tip tc cun gp bng cch no cho n nay cha bit y to nn mt dng nhim sc th co xon cc i nh c minh ha bng nh hin vi in t trn.Chiu rng ca nhim sc t khong 700nm. Cc gen nht nh lun c tm thy v tr c th ca chng trn nhim sc th k gia; iu ny cho thy: cc cp ng gi cao hn ca nhim sc th cng c tnh c hiu v chnh xc rt cao.

Chng 16

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Trong chu k t bo, cht nhim sc phi tri qua nhng thay i v cp ng gi ca n (xem Hnh 12.6). Khi nhum nhim sc th k trung gian v quan st di knh hin vi, cht nhim sc thng c thy dng khuch tn tng i u khp nhn t bo; lc ny, nhim sc th dng gin xon. Khi t bo chun b nguyn phn, cht nhim sc gp xon (c c), cui cng k gia hnh thnh nn mt s lng c trng cc nhim sc th c kch thc ngn v dy, c th phn bit c bng knh hin vi quang hc. Mc d cc nhim sc th k trung gian thng c mc c c thp hn nhiu so vi nhim sc th trong nguyn phn, nhng n cng c cc cp ng gi bc cao. Mt s cht nhim sc bao gm cc vng c cu trc si 10nm, bn cnh nhng vng khc ng gi thnh si 30nm; nhng vng ny c th tip tc ng gi thnh cc min "thng lng". Mc d mt nhim sc th k trung gian thng khng c mt khung nhim sc th r rt, nhng cc min thng lng ca n thng c biu hin nh kt vo mt lp mng bn trong mng nhn, v c l cng c th nh kt vo cc si cu trc nn mng li nhn. Bng cch nh kt nh vy, nhim sc th c t chc dng ph hp vi s biu hin ca cc gen. Cht nhim sc ca mi nhim sc th k trung gian chim mt khng gian c th trong nhn t bo, v cc si nhim sc thuc cc nhim sc th khc nhau khng vng mc vo nhau. Thm ch ngay trong k trung gian, cc cu trc ca nhim sc th bao gm tm ng v cc u mt cng nh mt s vng nhim sc th khc trong t bo tn ti trng thi co xon cao ging nh khi nhim sc th i vo k gia nguyn phn. Kiu cht nhim sc k trung gian quan st thy di knh hin vi dng kt c khng u n c c gi l d nhim sc, phn bit vi kiu cht nhim sc c mc kt c thp hn l nguyn nhim sc ("nhim sc tht"). Do c mc ng xon cao, cc b my biu hin thng tin di truyn ca t bo (nh b my phin m) kh c th tip cn ADN trong vng d nhim sc. Ngc li, s ng gi "lng lo" ca vng nguyn nhim sc cho php cc b my phin m c th tip cn c ADN; v vy, cc gen trong vng nguyn nhim sc c th c biu hin. Nhim sc th l mt cu trc nng ng; n c th c c, gin xon, bin i, thm ch thay i cu hnh theo yu cu ca cc qu trnh khc nhau trong t bo nh nguyn phn, gim phn v qu trnh biu hin ca cc gen. Hin nay, cc con ng iu ha s bin i ca cht nhim sc vn ang tip tc c cc nh khoa hc tp trung nghin cu. Tuy vy, mt vn tr nn r rng l cc histone khng ch n thun l cc ng ch c tnh tr ADN c th qun quanh trong cu trc cht nhim sc. Thay vo , s bin i ha hc ca cc protein histone c vai tr trc tip lm thay i mc t chc ca cht nhim sc v tham gia iu ha s biu hin ca cc gen. Terry Orr-Weaver, nh khoa hc c phng vn phn u ca khi kin thc Di truyn hc ny (cc trang 246 - 247), cng cc cng s ca mnh nghin cu v c ch phn t lin quan n ng hc nhim sc th trong nguyn phn v gim phn. Vi cc nghin cu Drosophila, cc nh khoa hc ch ra rng s phosphoryl ha mt s axit amin c th trong vng ui histone c vai tr quyt nh s ng thi ca nhim sc th trong k u ca gim phn I (Hnh 16.22).

Hnh 16.22

Nghin cu pht hin

S phosphoryl ha histone c vai tr g trong hot ng ca nhim sc th gim phn?

Th nghim Terry Orr-Weaver v cc cng s ti Vin Cng
ngh Massachusetts tin hnh gy t bin thc nghim rui gim nhm tm ra cc th t bin bt th vi suy ngh cho rng nhng th t bin c th lin quan n cc gen m ha cho cc protein gi vai tr quan trng trong nguyn phn. T , h tm ra mt t bin gen nhk-1 gy bt th rui ci. H bit rng sn phm ca gen ny l histone kinase-1 (NHK-1), mt enzym c vai tr phosphoryl ha mt axit amin c th vng ui ca histone H2A. H gi thit rng nguyn nhn gy nn tnh bt th l do enzym ny khng hot ng chc nng ng, dn n ng thi bt thng ca nhim sc th trong gim phn v qu trnh nguyn phn gp ri lon. kim tra gi thit, h quan st v so snh s vn ng ca nhim sc th trong gim phn cc t bo sinh trng ca rui t bin v rui kiu di (bnh thng). Trong mt th nghim, h dng mt thuc nhum hunh quang nh du ni nh v ca ADN v mt thuc nhum hunh quang xanh lc xc nh ni nh v ca protein condensin, l protein thng bao bc cc nhim sc th vo cui K u I v gip nhim sc th c c.

Kt qu Vo cui K u I, trong cc t bo sinh trng rui

kiu di, ADN v codensin nh v tp trung mt vng nh trong nhn (hnh di bn tri; mu vng l do thuc nhum v xanh c mt ng thi). Tuy vy, rui t bin, codensin khuch tn khp nhn; trong khi ADN ch tp trung vng bin quanh nhn (hnh di bn phi; mu ca ADN tng i m). Kt qu ny cho thy: codensin khng bao bc cc nhim sc th trong t bo ca cc rui t bin. Hu qu l cc nhim sc th khng c c c.

Codensin v ADN (mu vng)

Vin bao ngoi nhn

Codensin ADN (mu (mu xanh lc) vng quanh nhn)

Nhn t bo bnh thng Kt lun

Nhn t bo t bin

Do qu trnh gim phn khng th din ra bnh thng khi enzym histone kinase NHK-1 khng biu hin ng chc nng ca n, nn cc nh khoa hc kt lun rng s phosphoryl ha c th ui N ca histone H2A l thit yu hot ng ca cc nhim sc th trong gim phn c th din ra chnh xc.
histone code in meiosis: the histone kinase, NHK-1, is required for proper chromosomal architecture in Drosophila oocytes,Gene and Development 19: 2571 - 2582 (2005). iu g nu Gi s mt nh nghin cu tm ra rui gim mt th t bin mt axit amin vn bnh thng c phosphoryl ha bi histone kinase NHK-1. t bin ny s nh hng th no n hot ng ca nhim sc th trong gim phn cc t bo sinh trng?

Ngun I. Ivanovska, T. Khandan, T. Ito and T.L.Orr-Weaver, A

322

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Di truyn hc

S phosphoryl ha v cc bin i ha hc khc ca cc histone c nhiu tc ng n s hot ng ca cc gen, s c cp Chng 18. chng ny, chng ta tm hiu cc phn t ADN c t chc nh th no trong cc nhim sc th v bng cch no s sao chp ADN c th cung cp cc bn sao ca gen m b, m c th chuyn cho con con ci. Tuy vy, trong qu trnh di truyn, vic cc gen c sao chp v chuyn gia cc th h l cn thit nhng cha ; iu quan trng hn l cc thng tin di truyn phi c cc t bo s dng. Ni cch khc, cc gen phi dng "c biu hin". Trong chng tip theo, chng ta s xem cc t bo c th dch m thng tin di truyn c m ha trong cc phn t ADN nh th no.

Kim tra khi nim

16.3

ADN c bn t bo sinh vt nhn tht. 2. Hai thuc tnh gip phn bit d nhim sc v nguyn nhim sc l g?

1. Hy m t cu trc ca nucleosome, n v ng gi

3.

Mc d cc protein gip nhim sc th E. coli ng xon khng phi l histone, nhng theo bn thuc tnh no ging vi histone m cc protein ny cn phi c, xt v kh nng lin kt ADN?
Xem gi tr li Ph lc A.

iu g nu

Tng kt Chng
a phng tin Hy tham kho c s hc liu gm cc hnh nh ng ba chiu, cc bi hng dn dng file MP3, video, cc bi kim tra thc hnh, eBook v nhiu hc liu khc ti a ch Web www.masteringbio.com

Sao chp ADN: Quan st gn hn

ADN pol III tng hp mch dn u mt cch lin tc ADN m ADN pol III bt u tng hp ADN ti u 3 ca mi, ri ko di theo chiu 5 3 Mch ra chm c tng hp thnh nhng on Okazaki ngn ri c ni li bng ADN ligase

Tm tt cc khi nim chnh

Khi nim

16.1

ADN l vt cht di truyn (cc trang 305 310)

Tm kim vt cht di truyn: Qu trnh iu tra khoa hc Cc th nghim c tin hnh vi vi khun v phag cung cp cc bng chng thuyt phc u tin chng minh ADN l vt cht mang thng tin di truyn. Xy dng m hnh cu trc ADN: Qu trnh iu tra khoa hc Watson v Crick tm ra ADN c cu trc xon kp. Hai chui ng - phosphate i song song cun quanh pha ngoi phn t; cc baz nit hng vo pha trong, qua cc lin kt hydro chng kt cp c hiu vi nhau, gia A vi T v gia G vi C.

Primase tng hp mt on mi ARN ngn

c sa v sa cha ADN Cc enzym ADN polymerase c kh nng c sa mch ADN mi, thay th cc nucleotit sai hng. Trong c ch sa cha kt cp sai, cc enzym c th sa cha cc li tn ti sn. C ch sa cha bng ct b nucleotit l mt qu trnh c bn trong cc enzym c th ct b v thay th mt on di ADN mang cc nucleotit sai hng. Sao chp u tn cng ca phn t ADN u tn cng ca phn t ADN thuc nhim sc th sinh vt nhn tht thng ngn li sau mi chu k sao chp. S c mt ca u mt l trnh t lp li cc u tn cng ca cc phn t ADN mch thng l cch bo v cc gen gn u mt khi s "n mn". Enzym telomerase xc tc phn ng ko di u mt nhim sc th cc t bo mm sinh dc.
a phng tin

Cc baz nit Khung ng - phosphate

Lin kt hydro

a phng tin

Hot ng Th nghim Hershey - Chase Hot ng Cu trc ca ADN v ARN Hot ng Chui xon kp ADN
Khi nim

Hot ng Sao chp ADN: Tng quan iu tra M hnh sao chp ADN ng nh th no ? Hot ng Sao chp ADN: Quan st gn hn Hot ng Sao chp ADN: Tng kt
Khi nim

16.3

16.2

Nhiu protein phi hp trong sao chp v sa cha ADN (cc trang 311 319)
Nguyn l c bn: Kt cp baz vi mch lm khun Th nghim Meselson - Stahl cho thy ADN sao chp theo c ch bn bo ton: phn t m gin xon v mi mch ca n sau c dng lm khun tng hp nn cc mch mi trn c s nguyn tc kt cp b sung gia cc baz nit.

Mi nhim sc th gm mt phn t ADN c ng gi vi cc phn t protein (cc trang 320 323)

Nhim sc th vi khun thng l mt phn t ADN mch vng lin kt vi mt s protein. Cht nhim sc sinh vt nhn tht t hnh thnh nn nhim sc th gm ADN, cc histone v cc protein khc. Cc histone lin kt vi nhau v vi ADN hnh thnh nn nucleosome, l n v ng gi ADN c bn nht. Cc ui histone chc ra ngoi phn li nucleosome. S gp xon tip theo dn n s c c cc i ca cht nhim sc
Chng 16

C s di truyn hc phn t

323

 K gia. Trong cc t bo K trung gian, phn ln cht nhim sc trng thi c c thp hn (nguyn nhim sc), nhng mt s vng vn trng thi c c cao (d nhim sc). S bin i ca cc histone nh hng n s c c ca cht nhim sc.
a phng tin

6. S ko di mch dn u trong qu trnh sao chp ADN a. ngy cng ri xa chc sao chp. b. din ra theo chiu 3 5. c. to thnh cc on Okazaki d. ph thuc vo hot ng ca ADN polymerase e. khng cn mch lm khun 7. Khi t pht mt nhm amino, Adenine chuyn ha thnh Hypoxanthine, l mt baz him thng kt cp vi Thymine trong phn t ADN. S phi hp ca nhng phn t no c th sa cha c sai hng ny? a. nuclease, ADN polymerase, ADN ligase b. telomerase, primase, ADN polymerase c. telomerase, helicase, protein lin kt mch n d. ADN ligase, cc protein chc sao chp, adenylyl cyclase e. Nuclease, telomerase, primase 8. mi nucleosome, ADN c qun quanh bi a. cc polymerase d. mt phc kp thymine b. cc ribosome e. ADN vi v tinh c. cc histone Xem gi tr li Cc cu hi t nh gi Ph lc A.
web www.masteringbio.com lin h vi tin ha
a phng tin Thc hin bi Kim tra thc hnh ti trang

Hot ng S ng gi ADN
Kim tra kin thc ca bn

Cc cu hi t nh gi

1. Trong nghin cu ca mnh tin hnh chut v vi khun gy bnh lao phi, Griffith pht hin ra rng a. v protein t cc t bo gy bnh c kh nng chuyn ha cc t bo khng gy bnh thnh cc t bo gy bnh. b. cc t bo gy bnh sau khi un vn c kh nng gy bnh lao phi. c. mt s cht t cc t bo gy bnh c truyn sang cc t bo khng gy bnh v lm chng tr thnh dng gy bnh. d. v polysaccharide ca vi khun gy bnh lao phi. e. cc bacteriophag tim ADN vo vi khun. 2. Cc t bo E. coli c nui trong mi trng 15N, ri chuyn sang mi trng 14N v cho sinh trng qua hai th h (hai chu k sao chp ADN). Sau , ADN c tch chit t nhng t bo ny ri em li tm. Hy d on s phn b t trng ca ADN trong th nghim ny. a. Mt bng t trng cao v mt bng t trng thp b. Mt bng t trng trung bnh c. Mt bng t trng cao v mt bng t trng trung bnh d. Mt bng t trng thp v mt bng t trng trung bnh e. Mt bng t trng thp 3. Mt nh ha sinh hc phn lp v tinh sch c cc phn t cn thit cho qu trnh sao chp ADN. Khi c ta b sung thm ADN, s sao chp din ra, nhng mi phn t ADN bao gm mt mch bnh thng kt cp vi nhiu phn on ADN c chiu di gm vi trm nucleotit. Nhiu kh nng l c ta qun b sung vo hn hp thnh phn g? a. ADN polymerase d. Cc on Okazaki b. ADN ligase e. Primase c. Cc nucleotit 4. C s no dn n hin tng mch dn u v mch ra chm c tng hp khc nhau trong qu trnh sao chp ADN? a. im khi u sao chp ch c pha u 5. b. Enzym helicase v cc protein lin kt mch n ch hot ng u 5. c. ADN polymerase ch c th ni cc nucleotit mi vo pha u 3 ca mch ang ko di. d. ADN ligase ch hot ng theo chiu 5 3. e. Vo mi thi im, polymerase ch hot ng trn mt mch. 5. Khi phn tch thnh phn cc baz khc nhau trong mt mu ADN, kt qu no l ph hp vi nguyn tc b sung? a. A = G d. A = C b. A + G = C + T e. G = T c. A + T = G + T

9. Mt s vi khun c th p ng c vi cc tc nhn stress t mi trng bng vic tng tn s t bin trong qu trnh phn bo. Hin tng ny xy ra nh th no? N c u th g trong tin ha? Gii thch.
iu tra khoa hc

10.

v tip Xy dng cc m hnh l mt phng php quan trng trong cc nghin cu khoa hc. Hnh trn minh ha mt m hnh phc h sao chp ADN c m phng bi my tnh. Mch ADN gc v mch mi tng hp c phn bit bng cc mu khc nhau; tng t nh vy l ba loi protein: ADN pol III, protein cp trt v protein lin kt mch n. Trn c s kin thc hc c t chng ny, bn hy b sung cc thng tin lm r m hnh trn bng vic ch thch vo hnh tn cc mch ADN v mi loi protein, ng thi v thm mi tn ch r chiu m qu trnh sao chp ang din ra.

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