11/27/2013

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CHNG M U
GII THIU MN HC
ThS. Nguyn Thnh Lun
Email: luannt@cntp.edu.vn
KHOA CNSH & KTMT

BI GING CC PHNG PHP
NUI CY T BO
CNG MN HC
Tn hc phn: CC PHNG PHP NUI
CY T BO
Ma hc phn: 08220058
S tn ch: 2 (2, 0, 4)
Trinh : Dnh cho sinh vin i hc
H o to: i hc

Mc tiu mn hc
Trnh by c kin thc chung v nui cy t
bo, nhng hiu bit chung v mi trng v
iu kin nui cy;
Hiu r cc phng php nui cy c bn cc
m t bo khc nhau nh vi sinh vt, thc vt
hoc ng vt.
Hiu c s nh hng ca cc iu kin
ngoi cnh n kh nng nui cy v sinh
trng, pht trin ca t bo cng nh xut hin
cc bin d ca m hoc t bo sinh vt khi c
s thay i cc yu t ngoi cnh.
11/27/2013
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Ni dung c bn ca mn hc
C bn v nui cy t bo
Cu trc c bn ca t bo v s di
truyn ca n sau khi nui cy.
C ch iu ha v hot ng ca h
gen i vi t bo nui cy.
Cc phng php v k thut p dng
cho k thut nui cy t bo.
ng dng thc tin ca k thut nui cy
m & t bo.

Ti liu hc tp
Sch, gio trnh chnh
[1] Nguyn Hong Lc. Gio trnh Cng
ngh t bo, NXB i hc Hu. 2006
[2] Bi Trang Vit. Nui cy m v t bo
thc vt, Nh xut bn Gio dc. 2005
[3] Nguyn nh Giu, Nguyn Chi Mai,
Trn Th Vit Hng. Sinh l hc ngi v
ng vt. T sch i hc Khoa hc T
nhin TP HCM. 2000
Ti liu tham kho
Novakovic, G.V. & Freshney, R.I. Culture
of Specialized Cells Culture of Cells for
Tissue Engineering, Wiley-Liss, New
Jersey, USA. 2006
Mather, J.P. & Roberts, P.E. Introduction
to Cell and Tissue Culture: Theory And
Technique, Plenum Press, New York.
1998.

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Tiu ch nh gi mn hc
D lp: 80% tr ln tnh theo s tit
ln lp
Thi gia hc phn: 0%
Thi kt thc hc phn: 70%
Khc: 30% (Tho lun, tiu lun)

nh gia tiu lun (30%)
Seminar + tiu lun theo nhom
Thi gian: 15 pht/nhom, phi c y thnh
vin bo co
Ch v 1 ti liu chuyn ngnh v:
Nui cy t bo VSV
Nui cy t bo thc vt
Nui cy t bo ng vt
Ch trng vo cc phng php nui cy hin
i, ng dng cao
Mi nhom tiu lun: 4 ngi
BO CO TIU LUN
Sinh vin t chn lc tn ti ph hp
v cc lnh vc nui cy t bo vi sinh
vt, thc vt & ng vt.
Chn lc phng php nui cy hin i,
theo quy trnh nui cy theo cng nghip
(thnh phm).
c lng r rng gi c, phn tch u
v nhc im ca thit b v hiu sut
s dng cho i tng c th.
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Cu trc bi bo co seminar
Bao gm cc phn:
T VN
TNG QUAN
PHNG PHP NGHIN CU: ch trng
KT QU D KIN
THO LUN & KT LUN.
KIN NGH (NU C)

BO CO DI DNG FILE PPT C GHI
M
THI HN NP BI
LP TRNG TNG HP CC FILE
CA CC NHM VO A V GI A
V KHOA VO NGY 22/11/2013
(Khng nhn bi sau ngy trn)

BA A GHI: BI TIU LUN MN..
LP:.
Thi kt thc hc phn
Hnh thc thi: Trc nghim

Bao gm tt c ni dung c hc trong
ton b mn hc va gio trnh lin quan.
C phn cu hi ting Anh (nng cao)

Gm 4 phn chnh: i cng nui cy t
bo, PP nui cy TB VSV, Thc vt, ng
vt (C cu hi ting Anh)


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CNG CHI TIT
STT Ni dung S tit
1 Chng 1: M u 2
2 Chng 2: Sinh trng v bt ng ca t bo 4
3 Chng 3: ng hc sinh trng ca t bo 4
4 Chng 4: Thit k h ln men 4
5 Chng 5: Nui cy t bo vi sinh vt 4
6 Chng 6: Nui cy t bo ng vt 4
7 Chng 7: Nui cy t bo thc vt 4
8 Bao cao tiu lun 4
Cc ging vin phu trch mn hc
ThS. Nguyn Thnh Lun (PP nui cy
TB VSV - 3 tun u)
ThS. Phm Vn Lc (PP nui cy TB
thc vt - 2 tun tip theo)
ThS. Li inh Bin (PP nui cy TB
ng vt - 3 tun cui)
CHNG I: M U
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I CNG V CNG NGH T BO
Khi nim Cng ngh sinh hc hin i
Lch s hnh thnh Cng ngh t bo
Cc qu trnh sinh hc trong cng ngh
t bo
Ti sao li l: Nui cy t bo?
Cc u v nhc im ca cng ngh
t bo

Cng ngh sinh hc hin i
Cng ngh sinh hc hin i l cc qu
trnh sn xut quy m cng nghip m
nhn t tham gia trc tip v quyt nh l
cc t bo sng (vi sinh vt, thc vt v
ng vt). Mi t bo sng ca c th sinh
vt hot ng trong lnh vc sn xut ny
c xem nh mt l phn ng nh.
Cng ngh sinh hc hin i
Thut ng Cng ngh sinh
hc thng c s dng.
Vy CNSH l g?

Nhng k thut mi nh DNA
ti t hp (DNA recombinant)
v dung hp t bo (cell
fusion), c xem l lnh vc
cng ngh sinh hc hin i.
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Cu hi
Hy cho bit bn cht ca phng
php DNA ti t hp v dung hp t
bo?

Cc ng dng ca 2 phng php trn

Nhu cu ca CNSH hin i
Cc dc phm:
Sn xut Insulin cha bnh i tho
ng
Sn xut cc Hormone sinh trng ngi
iu tr bnh ci ca tr em
Cc Interferon (IFNs) chng vim nhim,
vaccine phng bnh v cc khng th n
dng dng chn on
QUY TRNH CNG
NGH NUI CY
T BO IN HNH
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Nhu cu ca CNSH hin i
Cc con ging sch bnh hoc kho mnh
hn, cc vt nui dng lm thc phm co
sn lng cao co th c pht trin
Cc loi cy trng quan trng c bin
i di truyn co cc tnh trng chng
chu stress
Chng chu cht dit c v khng cn
trng
Pht trin cc vi sinh vt c bin i di
truyn (genetically modification organisms)
Recombinant DNA (rDNA) - Cloning
L nhng k thut cho
php thao tc trc tip
nguyn liu di truyn
ca cc t bo ring
bit.
C th c s dng
pht trin cc vi
sinh vt sn xut cc
sn phm mi cng
nh cc c th hu
ch khc.
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Mc tiu chnh ca cng ngh rDNA
Gn mt gen ngoi lai (foreign gene) m
ha cho mt sn phm mong mun vo
trong cc dng DNA mch vng (plasmid
vector) ca vi khun

Chuyn gene vo trong mt c th vt
ch gen ngoi lai co th biu hin
sn xut sn phm ca no t c th ny.
Dung np t bo (cell fusion)
Dung np t bo
L qu trnh hnh thnh mt t bo lai n
(single hybrid cell) vi nhn v t bo cht
t hai loi t bo ring bit t hp cc
c im mong mun ca c hai loi t
bo ny.
ng dng trong vic ng dng sn xut
t bo lai trong sn xut khng th n
dng, ghep m t bo,
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Cng ngh nui cy t bo
Cc cng ngh DNA ti
t hp hoc dung np
t bo c khi u
bi nhng nghin cu
thun ty v cc kt
qu cui cng c th
pht trin thnh mt
loi t bo mi c th
sn xut sn phm c y
ngha thng mi
CNG NGH T BO
i hi phi pht trin thnh quy trnh cng
ngh vi cc cng ngh kh thi v co hiu
qu kinh t cao.

Nhu cu tm hiu cc phng php nui
cy t bo ti u nht.

Lch s hnh thnh
T khi no chng ta bit c cng ngh
nui cy t bo?

Cng ngh nui cy t bo ngy nay khc
vi cng ngh nui cy t bo ngy xa xa
nh th no?

Mc ch ca nui cy t bo hin i?
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Lch s hnh thnh
Mi c th sinh vt phc tp u gm nhiu
n v nh cc t bo hp thnh hay T
bo l n v sng cn bn ca mi cu trc
sinh vt (Schleiden v Schwann, 1838).
Lch s hnh thnh
Sydney Ringer (1836 1910),
ngi Anh khm ph ra dung
dch Ringers Solution quyt
nh nng u trng t cc
loi mui gm 6.5g NaCl, 0.42g
KCl, 0.25g CaCl
2
and 1 mole
Natri bicarbonate

ng dng trong tt c cc th
nghim in vitro v cc m v c
quan (c, tim ca chim, c,
ng vt c v..)
Lch s hnh thnh
Vo nm 1885, Roux
chuyn 1 mu a mi
trng nui cy t bo
thn kinh of 1 phi t
bo g v duy tr trong 1
mi trng mui saline,
m u cho vic pht
trin cng ngh nui cy
t bo.
Wilhelm Roux (1850 1924)
ngi c
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Lch s hnh thnh
T 19071910, sng
ch ra m t bo nhn
to t t bo thn kinh
ch trong mi trng
cha lymphocytes
Ross Granville Harrison
(M)
(1870 1959)
Lch s hnh thnh
n nm 1960,
Melcher v
Bergman l nhng
tc gi u tin tch
v nui cy thnh
cng t bo n
ca thc vt trong
cc bnh ln men
mi trng nhn
to.

Mc ch tim hiu
Cc cht xc tc sinh hc v t bo c
chn lc hoc sa i di truyn phi thch
hp cho cc hot ng sn xut
Ph hp vi m hnh sn xut quy m
cng nghip ng dng
Tm hiu cc phng php ti u nht
cho nhu cu thu nhn sinh khi 1 sn
phm thng mi
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Cc qu trnh sinh hc (Bio-processing)
Phi thu c cc cht xc tc sinh hc tt
nht (vi sinh vt, t bo ng vt, t bo thc
vt,hoc enzyme)
To ra mi trng tt nht co th cho s xc
tc bng cch thit k cc bioreactor/fermenter
thch hp vi phng thc ti u nht.
Phn tch cc sn phm mong mun t hn
hp phn ng trong mt phng thc kinh t
nht.
NHC LAI
Bio-prossessing
K thut to dng t bo
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B phn ng sinh hc
B phn ng sinh hc
Theo anh/ch, b phn ng sinh hc
(Bioreactor) l g?

Mc ch ca vic s dng b phn ng
sinh hc l g?
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Nui cy t bo
Nui cy t bo l
chn lc cc t bo
giai on pht
trin ti u nht
trong qu trnh sinh
trng v pht trin
ca t bo sinh vt
sng gm 4 giai
on l: thch nghi,
tng trng, cn
bng v suy tn.
Phng php nui cy t bo
im khc bit gia phng php nui
cy t bo n thc vt v ng vt,
VSV :
Mc v trng
Tn ti di dng huyn ph
Tc phn chia sinh khi
Phng thc sinh dng
Cc cht c to ra trong qu trnh trao
i cht
Cu trc c th sinh vt

Cc dng nui cy t bo
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Nui cy t bo vi sinh vt
Prokaryotes:
T bo vi khun
Virus v cc
bacteriophage

VSV Eukaryotes:
nm men, nm mc, to
(algae), V nguyn sinh
(protozoa)

S lc v lch s pht hin v gii thiu
khng sinh penicillin

Pht hin tnh c vo nm 1928 do A. Fleming


Khng sinh penicillin
Penicillin chrysogenum

Thuc h hiu kh
bt buc,
L loi nm si bo
t h.
Khi mi pht hin
trong mi trng
c, chng to ra hai
dng khun ty:
khun ty kh sinh v
khun ty dinh dng
mu trng
11/27/2013
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Da vo tnh cht ca canh trng
Nui cy chm

Nui b mt
Hiu qu kinh t
Vi ngun c cht chnh l glucose v ln
men theo phng php chm
Nng penicillin G trong dch ln men
nhng nm 80 - 90 ca th k XX t
khong 80.000 UI/ml (tng ng nng sut
khong 40 - 50 kg penicillin G/m
3
dch ln
men ).

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Nui cy t bo thc vt
Khi ngun t s tch
chit cc nhm m t r,
l, mm (explant culture)
thnh cc m so (callus)
trn mi trng dinh
dng in vitro
Nui cy m t bo thc vt
L s nui cy v
trng cc c quan, m,
t bo thc vt trn
mi trng nhn to
c kim sot.

Nui cy hoa lan Mokara
Hnh . Lan Mokara
Lan Mokara l mt loi
hoa p c gi tr kinh t
cao.
Mokara c hoa nhiu
mu sc c rt nhiu
ngi a thch & l loi
hoa c lng tiu th ln
trn th trng.
11/27/2013
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Nui cy t bo cn trng
Rui gim (Drosophila melanogaster)
Nghin cu phn ng sinh ha hc
siRNA (RNA thm lng)

Cc loi u trng ca bm, chu chu
v su
Recombinat protein
Xc nh cc nhm RNA hoc DNA kh
xc nh cc nhm sinh vt bc cao
Nui cy t bo ng vt
a dng:
Nui cy t bo gc
Sn xut vaccine
Bit ha dng t bo ng vt
Hot cht chng ung th
Khng th n dng
.
ng dng cc th nghim thnh cng
trn ngi phc v cho mc ch y hc
Cng ngh t bo gc (stem cell)
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Sn phm ca qu trnh sinh hc
Sn phm bin dng ca sinh vt
Sn phm s cp: carbohydrate, acid beo
(fatty acid), amino acid,
Sn phm th cp: alkaloid, nicotine,
berberin, ginsenosid, palitaxel,
bacteriocin,
Sn phm th cp gip p ng stress,
phng v chng vi sinh vt, cn trng,
ng dng trong tch chit hp cht sinh hc th cp
Hp cht ginsenosid
t cy Nhn sm
(Panax ginseng):
Tc dng n h thn
kinh trung ng
Gia tng sc khng
Tng cng co bp c
tim
H ng huyt

Saponin
Lm gim sc cng b mt, to bt nhiu khi
lc vi nc, co tc dng nh hoa v ty sch.
Lm v hng cu ngay nhng nng rt
long.
c vi c v ng vt mu lnh; co tc dng
dit nhng loi thn mm nh giun, sn, c
sn.
Kch ng nim mc, gy ht hi, mt.
C th to phc vi cholesterol hoc vi cc
cht 3--hydroxysteroid khc

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ng dng nui cy TB trong y dc
Bao gm:
Sn xut cc hot cht nh Diosgenin,
Codeine, Morphine, Atropine, Hyoscy-
amine, Scopolamine, Digoxin, Digitoxin,
Quinine, Reserpine, Artemisinin
Vaccine
Hormone tng trng/kch thch
Khng th n dng
T bo gc trong cy ghp m/t bo
Chn lc phng php lm tng sn lng
iu kin mi trng thch hp
Chn la chng co nng sut cao
Bin i tin cht
X ly vi cc cht cm ng
C nh t bo
Tit sn phm ra mi trng
Gy t bin t bo
Pht sinh hnh thi

C nh t bo
Mt h thng c nh co th duy tr s sng ca
t bo qua mt khong thi gian di v phng
thch phn ln sn phm ra mi trng ngoi
bo dng n nh:
S c nh thng b hn ch
Sinh khi ban u phi c nui cy dng
huyn ph t bo
Sn phm bt buc tit ra mi trng
Sn phm b thoi bin
Ngn cn s khuch tn ca t bo

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Tit sn phm ra mi trng
Lm tng kh nng sn xut nhng hp
cht th cp bng cch kch thch cho t
bo tit chng ra ngoi mi trng.
Tng tnh thm ca mng t bo lm
tng s tit sn phm ra mi trng
nhng kt qu t c vn rt hn ch
(thay i kh nng ca mng t bo)

Gy t bin t bo
Cc dng t bin iu ha v t bin khuyt dng c
s dng rng ri sn xut ra rt nhiu loi sn phm nh
amino acid, nucleotide, khng sinh,
S gia tng hot ng bin dng bng cch s dng cc t
bin iu ha co th thc hin
Vic chn lc cc kiu t bo ng dng cho mc ch nui
cy l mt yu t quan trng sn xut ra cc loi sn phm

Pht sinh hnh thi
Nui cy lng r (chuyn
plasmid Ri vo vt thng
trn m)
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Chn lc mi trng thch hp
iu kin mi trng thch hp
Thnh phn cc cht trong mi trng
nui cy, cht iu ha sinh trng thc
vt, pH, nhit , thng kh, lay
chuyn, nh sng,
V d: Auxin thng c b sung vo mi
trng cm ng s to m so, v no s
c gi li vi mt hm lng rt thp hoc
loi b hn t bo sn xut cc cht bin
dng

Cc lu trong s dng phng
php nui cy t bo
Khng nn nui cy t bo mt cch i tr m
khng tnh ton n kh nng to ra nhng sn
phm c bit ca loi o.
Khng phi tt c cc c quan u co kh nng
to ra cht m ta quan tm.
Trong qu trnh nui cy, lun lun phi tin hnh
tuyn chn nng cao nng sut to ra cc sn
phm bc 2.
B sung c cht tng ng.
Tm ra mi trng c hiu cho tng loi t bo

Cc u im
iu kin phn ng nh nhng, n ha:
qu trnh sinh hc t nh hng so vi qu
trnh ha hc
Tnh c hiu: 1 enzyme ch 1 hoc 2 c
cht xc tc
Tnh hiu lc: enzyme xc tc nhanh,
mnh so vi cht xc tc khc
Cc ti nguyn co th i mi
Cng ngh DNA ti t hp: h tr vic chn
lc dng/ging
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Cc u im
iu kin nui cy co th c kim sot
v co th c ti u cho vic sn xut
sn phm trao i cht th cp.
T bo co th c chn lc v ci thin
bng cch nhn dng in vitro
Co th d dng nghin cu chuyn hoa
cc cht trong t bo v c ch sn xut
sn phm trao i cht th cp.

Cc nhc im
Cc hn hp sn phm phc tp: xc
nh kh khn v kh tch chit
Cc mi trng nc loang: nh v
t tin
S nhim bn: d b ln t bi cc VSV
k sinh /cng sinh.
Khuynh hng hay bin i: do mi
trng hay t bin
KT THC CHNG I
THANK YOU FOR
YOUR ATTENTION!