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1
Examen de mi-session
CHMI 3216 – Bioingénierie de l’A.D.N.
Mardi, le 30 octobre 2007
Professeur : Eric R. Gauthier, Ph.D.
Votre nom : ___________________________________
Consignes :
1. Examen d’une durée de 85 minutes. 2. 6 questions, sur 13
pages, pour un total de 70 points. Compte pour 20% de la note
finale.
3. Répondre à toutes les questions, selon les instructions
fournies.
4. Inscrire vos réponses directement sur le questionnaire.
Limitez-vous à l’espace fourni (n’utilisez pas le verso des
pages).
5. Un document accompagnant le questionnaire vous fourni les
informations
nécessaires pour répondre aux questions 2.1, 3.1, 3.2 et 6
(cartes de restriction, liste d’enzymes, séquences, etc).
6. Note : toutes les séquences d’ADN sont présentées avec le
brin codant dans
l’orientation 5’ vers 3’ placée de gauche à droite.
7. L’utilisation des notes de cours et de livre est
interdite.
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Test de mi-session – CHMI 3216F – 30 octobre 2007
Question 1. Indiquez si les affirmations suivantes sont vraies
ou fausses (10 points - 2 points par question):
Enoncé Vrai ou Faux
1.1. L’ADN ligase du phage T4 requiert le NAD+ comme cofacteur
1.2. Les enzymes de restriction de type II sont des homodimères
1.3. L’activité 3’ 5’ exonucléase des ADN polymérases ne
fonctionne pas sur de l’ADN double brin.
1.4. Lors du PCR, une augmentation de la concentration d’ADN
polymérase diminue la fidélité de l’amplification.
1.5. Dans la préparation des banques d’ADNc, une des étapes
consiste à séparer les ADNc selon la quantité de radioactivité
incorporée lors de la transcription inverse.
Question 2. Répondez aux questions suivantes (12 points - 2
points par question):
Enoncé Réponse
2.1. Donnez la séquence d’une amorce permettant de séquencer un
ADNc cloné dans le site EcoR I du vecteur pEGFP-N1 (carte présentée
en annexe).
2.2. Ceci est le nom que l’on donne au phénomène selon lequel un
enzyme de restriction coupera des séquences d’ADN similaires, main
non-identique, à sa séquence cible lorsque la digestion enzymatique
est faite sous des conditions non-optimales.
2.3. Ceci est le type de séquence devant être présente en amont
(i.e. en 5’) d’un ADNc afin que ce dernier soit transcrit par une
ARN polymérase
2.4. Les bactériophages utilisés dans la préparation des banques
d’ADNc sont déficients dans le cycle……..
2.5. Dans la purification d’ARN total, quel est le rôle de la
guanidium thiocyanate?
2.6. Ceci est le substrat chromogène que l’on ajoute au milieu
de culture bactérien afin de détecter la présence d’activité
β-galactosidase.
2
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Test de mi-session – CHMI 3216F – 30 octobre 2007
Question 3. Donnez une réponse brève aux questions suivantes (18
points – 3 points par question): 3.1. Notre ami Stéphane (ben oui,
encore lui…) tente de cloner un fragment d’ADN de 450 pb
dans le vecteur pEGFP-N1 (un schéma de ce vecteur est montré en
annexe). Après avoir coupé son fragment d’ADN avec EcoR I et fait
la ligation dans le site EcoR I de pEGFP-N1, il transforme des
bactéries E. coli qu’il étale ensuite sur un milieu de culture
contenant de l’ampicilline. Lors de la transformation de ses
bactéries, Stéphane a la merveilleuse idée d’inclure plusieurs
échantillons contrôle. Il obtient les résultats suivants :
ADN ajouté aux bactéries lors de la transformation
Nombre de colonies par pétri
Contrôle : H2O 0 Contrôle : pBluescript > 1000 Contrôle :
pEGFP-N1 0 ADN/EcoRI + pEGFP-N1
0
Quel est la faille majeure dans le design expérimental qui
empêche Stéphane d’obtenir des bactéries E. coli contenant l’ADN
cloné dans pEGFP-N1? 3.2. Vous trouverez en annexe la séquence ADN
encodant la protéine p53. Quelle paire
d’amorce parmi les suivantes pourra être utilisée afin
d’amplifier le cadre de lecture ouvert (open reading frame - ORF)
complet de p53?
a) amorce 5’ : atggaggagccgcagtcag amorce 3’:
agggcctgactcagactga b) amorce 5’ : ctgactgcggctcctccat amorce 3’ :
tcagtctgagtcaggccct c) amorce 5’ : tcctcggcgtcagtcatgg amorce 3’ :
agggcctgactcagactga d) amorce 5’ : atggaggagccgcagtcag amorce 3’ :
tcagtctgagtcaggccct
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Test de mi-session – CHMI 3216F – 30 octobre 2007
3.3. Expliquez le principe de la méthode « touchdown PCR ». Quel
est l’avantage principal de cette approche?
3.4. Expliquez pourquoi il est important d’exprimer les ADNc
sous forme de protéine de fusion
avec la β-galactosidase lors du criblage d’une banque d’ADNc
(clonée dans le vecteur λgt11) avec un anticorps?
4
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Test de mi-session – CHMI 3216F – 30 octobre 2007
3.5. Notre ami Chris prépare une banque d’ADNc. Malheureusement
pour lui, l’analyse
aléatoire (i.e. au hasard) de plages de lyse révèle que la très
grande majorité des ADNc clonés dans le phage λ sont de petite
taille (en dessous de 2 kpb). Donnez une cause probable de ce
problème, et expliquez comment Chris pourrait résoudre cette
difficulté.
3.6. Expliquez comment on devra procéder afin de marquer
radioactivement l’extrémité 5’
d’une courte séquence d’ADN simple brin.
5
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Test de mi-session – CHMI 3216F – 30 octobre 2007
Question 4. (5 points). Décrivez les différentes étapes requises
lors de l’isolation de plasmides à partir de colonies
bactériennes.
6
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Test de mi-session – CHMI 3216F – 30 octobre 2007
Question 5 (15 points). Expliquez en détail les étapes de la
préparation et du criblage (avec un ADNc de 400pb) d’une banque
d’ADNc à partir de la lignée cellulaire HepG2 (cellules de foie
humain).
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Test de mi-session – CHMI 3216F – 30 octobre 2007
Question 5 (suite)
8
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Test de mi-session – CHMI 3216F – 30 octobre 2007
Question 5 (suite)
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Test de mi-session – CHMI 3216F – 30 octobre 2007
Question 6 (15 points). On vous donne un fragment d’ADN
correspondant au cadre de lecture ouvert complet de eIF2α. Préparez
une stratégie de clonage de cet ADN dans le MCS de pOPRSV/MCS.
Indiquez comment vous utiliseriez des enzymes de restriction afin
de confirmer l’orientation de ce fragment dans pOPRSV. NOTE 1:
Votre réponse doit inclure tous les réactifs (enzymes, cofacteurs,
substrats) que vous utiliserez pour cette expérience. NOTE 2: le
cadre de lecture ouvert complet (et seulement le cadre de lecture
ouvert) doit être inséré dans le vecteur de manière directionnelle,
de telle sorte que le brin codant de eIF2α soit dans la bonne
orientation pour être transcrit avec la polymérase T7. Vous
trouverez en annexe la séquence de l’ADNc encodant la protéine
eIF2α, ainsi que sa carte de restriction. Vous trouverez aussi la
carte de restriction du vecteur pOPRSV1/MCS. NOTE 3.
Malheureusement, votre copain Eric (je vous laisse deviner lequel…)
a brisé votre seul et unique « thermocycler ». Donc, vous ne pouvez
pas utiliser le PCR pour cette expérience.
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Test de mi-session – CHMI 3216F – 30 octobre 2007
Question 6 (suite)
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Test de mi-session – CHMI 3216F – 30 octobre 2007
Question 6 (suite)
12
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Test de mi-session – CHMI 3216F – 30 octobre 2007
Question 6 (suite)
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CHMI 3216 F
Bioingénierie de l’ADN
Annexe au Test de mi-session
30 octobre 2007
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Question 2.1 et 3.1
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Question 3.2 LOCUS NM_000546 2629 bp mRNA linear PRI 24-DEC-2006
DEFINITION Homo sapiens tumor protein p53 (Li-Fraumeni syndrome)
(TP53), mRNA. ACCESSION NM_000546 VERSION NM_000546.2 GI:8400737
KEYWORDS . SOURCE Homo sapiens (human) ORGANISM Homo sapiens
Eukaryota; Metazoa; Chordata; Craniata; Vertebrata; Euteleostomi;
Mammalia; Eutheria; Euarchontoglires; Primates; Haplorrhini;
Catarrhini; Hominidae; Homo. REFERENCE 1 (bases 1 to 2629) AUTHORS
Zeng,P.Y. and Berger,S.L. TITLE LKB1 is recruited to the p21/WAF1
promoter by p53 to mediate transcriptional activation JOURNAL
Cancer Res. 66 (22), 10701-10708 (2006) PUBMED 17108107 REMARK
GeneRIF: Chromatin immunoprecipitation analyses show that LKB1 is
recruited directly to the p21/WAF1 promoter, as well as to other
p53 activated promoters, in a p53-dependent fashion. REFERENCE 2
(bases 1 to 2629) AUTHORS Laurie,N.A., Donovan,S.L., Shih,C.S.,
Zhang,J., Mills,N., Fuller,C., Teunisse,A., Lam,S., Ramos,Y.,
Mohan,A., Johnson,D., Wilson,M., Rodriguez-Galindo,C., Quarto,M.,
Francoz,S., Mendrysa,S.M., Guy,R.K., Marine,J.C., Jochemsen,A.G.
and Dyer,M.A. TITLE Inactivation of the p53 pathway in
retinoblastoma JOURNAL Nature 444 (7115), 61-66 (2006) PUBMED
17080083 REMARK GeneRIF: data provide evidence that the p53 pathway
is inactivated in retinoblastoma and that this cancer does not
originate from intrinsically death-resistant cells as previously
thought REFERENCE 3 (bases 1 to 2629) AUTHORS Cai,Q.L.,
Knight,J.S., Verma,S.C., Zald,P. and Robertson,E.S. TITLE EC5S
ubiquitin complex is recruited by KSHV latent antigen LANA for
degradation of the VHL and p53 tumor suppressors JOURNAL PLoS
Pathog. 2 (10), E116 (2006) PUBMED 17069461 REMARK GeneRIF: In
transfected cells and KSHV-infected B lymphoma cells, KSHV-encoded
latency-associated nuclear antigen (LANA) expression stimulates
degradation of tumor suppressors von Hippel-Lindau and p53.
REFERENCE 4 (bases 1 to 2629) AUTHORS Feuerborn,A., Moritz,C., Von
Bonin,F., Dobbelstein,M., Trumper,L., Sturzenhofecker,B. and
Kube,D. TITLE Dysfunctional p53 deletion mutants in cell lines
derived from Hodgkin's lymphoma JOURNAL Leuk. Lymphoma 47 (9),
1932-1940 (2006) PUBMED 17065008
http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?id=9606http://www.ncbi.nlm.nih.gov/sites/entrez?cmd=Retrieve&db=pubmed&list_uids=17108107http://www.ncbi.nlm.nih.gov/sites/entrez?cmd=Retrieve&db=pubmed&list_uids=17080083http://www.ncbi.nlm.nih.gov/sites/entrez?cmd=Retrieve&db=pubmed&list_uids=17069461http://www.ncbi.nlm.nih.gov/sites/entrez?cmd=Retrieve&db=pubmed&list_uids=17065008
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REFERENCE 5 (bases 1 to 2629) AUTHORS Perra,M.T., Maxia,C.,
Corbu,A., Minerba,L., Demurtas,P., Colombari,R., Murtas,D.,
Bravo,S., Piras,F. and Sirigu,P. TITLE Oxidative stress in
pterygium: relationship between p53 and 8-hydroxydeoxyguanosine
JOURNAL (er) Mol. Vis. 12, 1136-1142 (2006) PUBMED 17093398 REMARK
GeneRIF: Although pterygium has limited local invasion and
ainability to metastasize, concomitant presence of altered p53 in
8-OHdG-immunoreactive cells could provide evidence of apparent
genetic instability, which is in contrast to its benign clinical
course. REFERENCE 6 (bases 1 to 2629) AUTHORS Seto,E., Usheva,A.,
Zambetti,G.P., Momand,J., Horikoshi,N., Weinmann,R., Levine,A.J.
and Shenk,T. TITLE Wild-type p53 binds to the TATA-binding protein
and represses transcription JOURNAL Proc. Natl. Acad. Sci. U.S.A.
89 (24), 12028-12032 (1992) PUBMED 1465435REFERENCE 7 (bases 1 to
2629) AUTHORS Baudier,J., Delphin,C., Grunwald,D., Khochbin,S. and
Lawrence,J.J. TITLE Characterization of the tumor suppressor
protein p53 as a protein kinase C substrate and a S100b-binding
protein JOURNAL Proc. Natl. Acad. Sci. U.S.A. 89 (23), 11627-11631
(1992) PUBMED 1454855REFERENCE 8 (bases 1 to 2629) AUTHORS
Crook,T., Wrede,D., Tidy,J.A., Mason,W.P., Evans,D.J. and
Vousden,K.H. TITLE Clonal p53 mutation in primary cervical cancer:
association with human-papillomavirus-negative tumours JOURNAL
Lancet 339 (8801), 1070-1073 (1992) PUBMED 1349102REFERENCE 9
(bases 1 to 2629) AUTHORS Iavarone,A., Matthay,K.K.,
Steinkirchner,T.M. and Israel,M.A. TITLE Germ-line and somatic p53
gene mutations in multifocal osteogenic sarcoma JOURNAL Proc. Natl.
Acad. Sci. U.S.A. 89 (9), 4207-4209 (1992) PUBMED 1349175REFERENCE
10 (bases 1 to 2629) AUTHORS Zhukov-Verezhnikov,N.N.,
Anisimov,P.I., Goncharova,N.S., Bochko,G.M. and Karaseva,Z.N. TITLE
[Study of the heterogenetic antigens in vaccinal preparations of V.
cholerae] JOURNAL Biull Eksp Biol Med 82 (8), 961-962 (1976) PUBMED
1088347COMMENT REVIEWED REFSEQ: This record has been curated by
NCBI staff. The reference sequence was derived from M13121.1,
M22881.1, U94788.1 and X02469.1. [WARNING] On Dec 29, 2006 this
sequence was replaced by gi:120407067. On Jun 9, 2000 this sequence
version replaced gi:4507636. Summary: Tumor protein p53, a nuclear
protein, plays an essential role in the regulation of cell cycle,
specifically in the
http://www.ncbi.nlm.nih.gov/sites/entrez?cmd=Retrieve&db=pubmed&list_uids=17093398http://www.ncbi.nlm.nih.gov/sites/entrez?cmd=Retrieve&db=pubmed&list_uids=1465435http://www.ncbi.nlm.nih.gov/sites/entrez?cmd=Retrieve&db=pubmed&list_uids=1454855http://www.ncbi.nlm.nih.gov/sites/entrez?cmd=Retrieve&db=pubmed&list_uids=1349102http://www.ncbi.nlm.nih.gov/sites/entrez?cmd=Retrieve&db=pubmed&list_uids=1349175http://www.ncbi.nlm.nih.gov/sites/entrez?cmd=Retrieve&db=pubmed&list_uids=1088347http://www.ncbi.nlm.nih.gov/RefSeq/http://www.ncbi.nlm.nih.gov/entrez/viewer.fcgi?val=M13121.1http://www.ncbi.nlm.nih.gov/entrez/viewer.fcgi?val=M22881.1http://www.ncbi.nlm.nih.gov/entrez/viewer.fcgi?val=U94788.1http://www.ncbi.nlm.nih.gov/entrez/viewer.fcgi?val=X02469.1http://www.ncbi.nlm.nih.gov/entrez/viewer.fcgi?val=120407067http://www.ncbi.nlm.nih.gov/entrez/viewer.fcgi?val=4507636
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transition from G0 to G1. It is found in very low levels in
normal cells, however, in a variety of transformed cell lines, it
is expressed in high amounts, and believed to contribute to
transformation and malignancy. p53 is a DNA-binding protein
containing DNA-binding, oligomerization and transcription
activation domains. It is postulated to bind as a tetramer to a
p53-binding site and activate expression of downstream genes that
inhibit growth and/or invasion, and thus function as a tumor
suppressor. Mutants of p53 that frequently occur in a number of
different human cancers fail to bind the consensus DNA binding
site, and hence cause the loss of tumor suppressor activity.
Alterations of the TP53 gene occur not only as somatic mutations
in human malignancies, but also as germline mutations in some
cancer-prone families with Li-Fraumeni syndrome. Publication
Note: This RefSeq record includes a subset of the publications that
are available for this gene. Please see the Entrez Gene record to
access additional publications. COMPLETENESS: full length. FEATURES
Location/Qualifiers source 1..2629 /organism="Homo sapiens"
/mol_type="mRNA" /db_xref="taxon:9606" /chromosome="17"
/map="17p13.1" gene 1..2629 /gene="TP53" /note="tumor protein p53
(Li-Fraumeni syndrome); synonyms: p53, LFS1, TRP53"
/db_xref="GeneID:7157" /db_xref="HGNC:11998" /db_xref="HPRD:01859"
/db_xref="MIM:191170" CDS 252..1433 /gene="TP53"
/GO_component="cytoplasm [PMID 7720704]; insoluble fraction [PMID
12915590]; mitochondrion [PMID 12667443]; nuclear matrix [PMID
11080164]; nucleolus [PMID 12080348]; nucleoplasm [PMID 11080164]
[PMID 12915590]; nucleus [PMID 7720704]" /GO_function="ATP binding
[PMID 8183576]; copper ion binding [PMID 7824276]; DNA strand
annealing activity [PMID 8183576]; enzyme binding [PMID 10666337]
[PMID 15577914]; metal ion binding; nuclease activity [PMID
11002423]; protein binding [PMID 15782130]; protein
heterodimerization activity [PMID 10837489]; protein N-terminus
binding [PMID 11861836]; transcription factor activity [PMID
7587074]; zinc ion binding [PMID 10065153]" /GO_process="apoptosis
[PMID 7720704]; base-excision
http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?id=9606http://www.ncbi.nlm.nih.gov/entrez/viewer.fcgi?val=8400737&from=1&to=2629&view=gbwithpartshttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=gene&cmd=Retrieve&dopt=full_report&list_uids=7157http://www.genenames.org/data/hgnc_data.php?hgnc_id=11998http://www.hprd.org/protein/01859http://www.ncbi.nlm.nih.gov/entrez/dispomim.cgi?id=191170http://www.ncbi.nlm.nih.gov/entrez/viewer.fcgi?val=8400737&from=252&to=1433&view=gbwithpartshttp://amigo.geneontology.org/cgi-bin/amigo/go.cgi?view=details&depth=1&query=0005737http://www.ncbi.nlm.nih.gov/sites/entrez?cmd=Retrieve&db=pubmed&list_uids=7720704http://amigo.geneontology.org/cgi-bin/amigo/go.cgi?view=details&depth=1&query=0005626http://amigo.geneontology.org/cgi-bin/amigo/go.cgi?view=details&depth=1&query=0005626http://www.ncbi.nlm.nih.gov/sites/entrez?cmd=Retrieve&db=pubmed&list_uids=12915590http://amigo.geneontology.org/cgi-bin/amigo/go.cgi?view=details&depth=1&query=0005739http://www.ncbi.nlm.nih.gov/sites/entrez?cmd=Retrieve&db=pubmed&list_uids=12667443http://amigo.geneontology.org/cgi-bin/amigo/go.cgi?view=details&depth=1&query=0016363http://www.ncbi.nlm.nih.gov/sites/entrez?cmd=Retrieve&db=pubmed&list_uids=11080164http://amigo.geneontology.org/cgi-bin/amigo/go.cgi?view=details&depth=1&query=0005730http://www.ncbi.nlm.nih.gov/sites/entrez?cmd=Retrieve&db=pubmed&list_uids=12080348http://amigo.geneontology.org/cgi-bin/amigo/go.cgi?view=details&depth=1&query=0005654http://www.ncbi.nlm.nih.gov/sites/entrez?cmd=Retrieve&db=pubmed&list_uids=11080164http://www.ncbi.nlm.nih.gov/sites/entrez?cmd=Retrieve&db=pubmed&list_uids=12915590http://amigo.geneontology.org/cgi-bin/amigo/go.cgi?view=details&depth=1&query=0005634http://www.ncbi.nlm.nih.gov/sites/entrez?cmd=Retrieve&db=pubmed&list_uids=7720704http://amigo.geneontology.org/cgi-bin/amigo/go.cgi?view=details&depth=1&query=0005524http://www.ncbi.nlm.nih.gov/sites/entrez?cmd=Retrieve&db=pubmed&list_uids=8183576http://amigo.geneontology.org/cgi-bin/amigo/go.cgi?view=details&depth=1&query=0005507http://amigo.geneontology.org/cgi-bin/amigo/go.cgi?view=details&depth=1&query=0005507http://www.ncbi.nlm.nih.gov/sites/entrez?cmd=Retrieve&db=pubmed&list_uids=7824276http://amigo.geneontology.org/cgi-bin/amigo/go.cgi?view=details&depth=1&query=0000739http://www.ncbi.nlm.nih.gov/sites/entrez?cmd=Retrieve&db=pubmed&list_uids=8183576http://amigo.geneontology.org/cgi-bin/amigo/go.cgi?view=details&depth=1&query=0019899http://www.ncbi.nlm.nih.gov/sites/entrez?cmd=Retrieve&db=pubmed&list_uids=10666337http://www.ncbi.nlm.nih.gov/sites/entrez?cmd=Retrieve&db=pubmed&list_uids=15577914http://amigo.geneontology.org/cgi-bin/amigo/go.cgi?view=details&depth=1&query=0046872http://amigo.geneontology.org/cgi-bin/amigo/go.cgi?view=details&depth=1&query=0004518http://www.ncbi.nlm.nih.gov/sites/entrez?cmd=Retrieve&db=pubmed&list_uids=11002423http://amigo.geneontology.org/cgi-bin/amigo/go.cgi?view=details&depth=1&query=0005515http://www.ncbi.nlm.nih.gov/sites/entrez?cmd=Retrieve&db=pubmed&list_uids=15782130http://amigo.geneontology.org/cgi-bin/amigo/go.cgi?view=details&depth=1&query=0046982http://amigo.geneontology.org/cgi-bin/amigo/go.cgi?view=details&depth=1&query=0046982http://www.ncbi.nlm.nih.gov/sites/entrez?cmd=Retrieve&db=pubmed&list_uids=10837489http://amigo.geneontology.org/cgi-bin/amigo/go.cgi?view=details&depth=1&query=0047485http://amigo.geneontology.org/cgi-bin/amigo/go.cgi?view=details&depth=1&query=0047485http://www.ncbi.nlm.nih.gov/sites/entrez?cmd=Retrieve&db=pubmed&list_uids=11861836http://amigo.geneontology.org/cgi-bin/amigo/go.cgi?view=details&depth=1&query=0003700http://amigo.geneontology.org/cgi-bin/amigo/go.cgi?view=details&depth=1&query=0003700http://www.ncbi.nlm.nih.gov/sites/entrez?cmd=Retrieve&db=pubmed&list_uids=7587074http://amigo.geneontology.org/cgi-bin/amigo/go.cgi?view=details&depth=1&query=0008270http://www.ncbi.nlm.nih.gov/sites/entrez?cmd=Retrieve&db=pubmed&list_uids=10065153http://amigo.geneontology.org/cgi-bin/amigo/go.cgi?view=details&depth=1&query=0006915http://www.ncbi.nlm.nih.gov/sites/entrez?cmd=Retrieve&db=pubmed&list_uids=7720704http://amigo.geneontology.org/cgi-bin/amigo/go.cgi?view=details&depth=1&query=0006284
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repair [PMID 15116721]; caspase activation via cytochrome c
[PMID 12667443]; cell aging [PMID 12080348]; cell cycle; cell cycle
arrest [PMID 8675009]; cell differentiation [PMID 10065150]; cell
proliferation [PMID 10065150]; DNA damage response, signal
transduction by p53 class mediator resulting in induction of
apoptosis [PMID 14654789]; negative regulation of cell growth [PMID
8986812]; negative regulation of helicase activity [PMID 7663514];
nucleotide-excision repair [PMID 7663514]; protein complex assembly
[PMID 12915590]; protein tetramerization [PMID 15116721];
regulation of mitochondrial membrane permeability [PMID 15116721];
regulation of transcription, DNA-dependent [PMID 7587074]; response
to DNA damage stimulus [PMID 8986812]; transcription" /note="p53
tumor suppressor" /codon_start=1 /product="tumor protein p53"
/protein_id="NP_000537.2" /db_xref="GI:8400738"
/db_xref="CCDS:CCDS11118.1" /db_xref="GeneID:7157"
/db_xref="HGNC:11998" /db_xref="HPRD:01859" /db_xref="MIM:191170"
/translation="MEEPQSDPSVEPPLSQETFSDLWKLLPENNVLSPLPSQAMDDLM
LSPDDIEQWFTEDPGPDEAPRMPEAAPRVAPAPAAPTPAAPAPAPSWPLSSSVPSQKT
YQGSYGFRLGFLHSGTAKSVTCTYSPALNKMFCQLAKTCPVQLWVDSTPPPGTRVRAM
AIYKQSQHMTEVVRRCPHHERCSDSDGLAPPQHLIRVEGNLRVEYLDDRNTFRHSVVV
PYEPPEVGSDCTTIHYNYMCNSSCMGGMNRRPILTIITLEDSSGNLLGRNSFEVRVCA
CPGRDRRTEEENLRKKGEPHHELPPGSTKRALPNNTSSSPQPKKKPLDGEYFTLQIRG
RERFEMFRELNEALELKDAQAGKEPGGSRAHSSHLKSKKGQSTSRHKKLMFKTEGPDS D"
misc_feature 465..467 /gene="TP53" /note="A single base change (g
or c at nt 466) gives rise to two variants with either R (more
frequent) or P at aa position 72; this aa change is responsible for
the observed difference in electrophoretic mobility between the two
variants; Region: polymorphic variant" misc_feature 1185..1220
/gene="TP53" /note="Region: Nuclear localization signal"
misc_feature 1194..1196 /gene="TP53"
http://amigo.geneontology.org/cgi-bin/amigo/go.cgi?view=details&depth=1&query=0006284http://www.ncbi.nlm.nih.gov/sites/entrez?cmd=Retrieve&db=pubmed&list_uids=15116721http://amigo.geneontology.org/cgi-bin/amigo/go.cgi?view=details&depth=1&query=0008635http://amigo.geneontology.org/cgi-bin/amigo/go.cgi?view=details&depth=1&query=0008635http://amigo.geneontology.org/cgi-bin/amigo/go.cgi?view=details&depth=1&query=0008635http://www.ncbi.nlm.nih.gov/sites/entrez?cmd=Retrieve&db=pubmed&list_uids=12667443http://amigo.geneontology.org/cgi-bin/amigo/go.cgi?view=details&depth=1&query=0007569http://www.ncbi.nlm.nih.gov/sites/entrez?cmd=Retrieve&db=pubmed&list_uids=12080348http://amigo.geneontology.org/cgi-bin/amigo/go.cgi?view=details&depth=1&query=0007049http://amigo.geneontology.org/cgi-bin/amigo/go.cgi?view=details&depth=1&query=0007049http://amigo.geneontology.org/cgi-bin/amigo/go.cgi?view=details&depth=1&query=0007050http://www.ncbi.nlm.nih.gov/sites/entrez?cmd=Retrieve&db=pubmed&list_uids=8675009http://amigo.geneontology.org/cgi-bin/amigo/go.cgi?view=details&depth=1&query=0030154http://www.ncbi.nlm.nih.gov/sites/entrez?cmd=Retrieve&db=pubmed&list_uids=10065150http://amigo.geneontology.org/cgi-bin/amigo/go.cgi?view=details&depth=1&query=0008283http://www.ncbi.nlm.nih.gov/sites/entrez?cmd=Retrieve&db=pubmed&list_uids=10065150http://amigo.geneontology.org/cgi-bin/amigo/go.cgi?view=details&depth=1&query=0042771http://amigo.geneontology.org/cgi-bin/amigo/go.cgi?view=details&depth=1&query=0042771http://amigo.geneontology.org/cgi-bin/amigo/go.cgi?view=details&depth=1&query=0042771http://amigo.geneontology.org/cgi-bin/amigo/go.cgi?view=details&depth=1&query=0042771http://www.ncbi.nlm.nih.gov/sites/entrez?cmd=Retrieve&db=pubmed&list_uids=14654789http://amigo.geneontology.org/cgi-bin/amigo/go.cgi?view=details&depth=1&query=0030308http://www.ncbi.nlm.nih.gov/sites/entrez?cmd=Retrieve&db=pubmed&list_uids=8986812http://amigo.geneontology.org/cgi-bin/amigo/go.cgi?view=details&depth=1&query=0051097http://www.ncbi.nlm.nih.gov/sites/entrez?cmd=Retrieve&db=pubmed&list_uids=7663514http://amigo.geneontology.org/cgi-bin/amigo/go.cgi?view=details&depth=1&query=0006289http://www.ncbi.nlm.nih.gov/sites/entrez?cmd=Retrieve&db=pubmed&list_uids=7663514http://amigo.geneontology.org/cgi-bin/amigo/go.cgi?view=details&depth=1&query=0006461http://amigo.geneontology.org/cgi-bin/amigo/go.cgi?view=details&depth=1&query=0006461http://amigo.geneontology.org/cgi-bin/amigo/go.cgi?view=details&depth=1&query=0006461http://www.ncbi.nlm.nih.gov/sites/entrez?cmd=Retrieve&db=pubmed&list_uids=12915590http://amigo.geneontology.org/cgi-bin/amigo/go.cgi?view=details&depth=1&query=0051262http://www.ncbi.nlm.nih.gov/sites/entrez?cmd=Retrieve&db=pubmed&list_uids=15116721http://amigo.geneontology.org/cgi-bin/amigo/go.cgi?view=details&depth=1&query=0046902http://amigo.geneontology.org/cgi-bin/amigo/go.cgi?view=details&depth=1&query=0046902http://www.ncbi.nlm.nih.gov/sites/entrez?cmd=Retrieve&db=pubmed&list_uids=15116721http://amigo.geneontology.org/cgi-bin/amigo/go.cgi?view=details&depth=1&query=0006355http://amigo.geneontology.org/cgi-bin/amigo/go.cgi?view=details&depth=1&query=0006355http://amigo.geneontology.org/cgi-bin/amigo/go.cgi?view=details&depth=1&query=0006355http://www.ncbi.nlm.nih.gov/sites/entrez?cmd=Retrieve&db=pubmed&list_uids=7587074http://amigo.geneontology.org/cgi-bin/amigo/go.cgi?view=details&depth=1&query=0006974http://amigo.geneontology.org/cgi-bin/amigo/go.cgi?view=details&depth=1&query=0006974http://www.ncbi.nlm.nih.gov/sites/entrez?cmd=Retrieve&db=pubmed&list_uids=8986812http://amigo.geneontology.org/cgi-bin/amigo/go.cgi?view=details&depth=1&query=0006350http://www.ncbi.nlm.nih.gov/entrez/viewer.fcgi?val=NP_000537.2http://www.ncbi.nlm.nih.gov/CCDS/CcdsBrowse.cgi?REQUEST=CCDS&DATA=CCDS11118.1http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=gene&cmd=Retrieve&dopt=full_report&list_uids=7157http://www.genenames.org/data/hgnc_data.php?hgnc_id=11998http://www.hprd.org/protein/01859http://www.ncbi.nlm.nih.gov/entrez/dispomim.cgi?id=191170http://www.ncbi.nlm.nih.gov/entrez/viewer.fcgi?val=8400738&itemID=5&view=gpwithpartshttp://www.ncbi.nlm.nih.gov/entrez/viewer.fcgi?val=8400738&from=312&to=323&view=gpwithpartshttp://www.ncbi.nlm.nih.gov/entrez/viewer.fcgi?val=8400738&itemID=7&view=gpwithparts
-
/note="phosphorylation site" STS 354..552 /gene="TP53"
/standard_name="GDB:177814" /db_xref="UniSTS:154969" STS 368..626
/gene="TP53" /standard_name="GDB:177797" /db_xref="UniSTS:154963"
STS 729..803 /gene="TP53" /standard_name="PMC340938P3"
/db_xref="UniSTS:273171" STS 1004..1202 /gene="TP53"
/standard_name="PMC310707P2" /db_xref="UniSTS:272633" STS
1008..1082 /gene="TP53" /standard_name="GDB:190076"
/db_xref="UniSTS:155620" STS 1503..1619 /gene="TP53"
/standard_name="D17S1678" /db_xref="UniSTS:82485" STS 2503..2579
/gene="TP53" /standard_name="WI-20715" /db_xref="UniSTS:59997"
polyA_signal 2612..2617 /gene="TP53" polyA_site 2629 /gene="TP53"
ORIGIN 1 acttgtcatg gcgactgtcc agctttgtgc caggagcctc gcaggggttg
atgggattgg 61 ggttttcccc tcccatgtgc tcaagactgg cgctaaaagt
tttgagcttc tcaaaagtct 121 agagccaccg tccagggagc aggtagctgc
tgggctccgg ggacactttg cgttcgggct 181 gggagcgtgc tttccacgac
ggtgacacgc ttccctggat tggcagccag actgccttcc 241 gggtcactgc
catggaggag ccgcagtcag atcctagcgt cgagccccct ctgagtcagg 301
aaacattttc agacctatgg aaactacttc ctgaaaacaa cgttctgtcc cccttgccgt
361 cccaagcaat ggatgatttg atgctgtccc cggacgatat tgaacaatgg
ttcactgaag 421 acccaggtcc agatgaagct cccagaatgc cagaggctgc
tccccgcgtg gcccctgcac 481 cagcagctcc tacaccggcg gcccctgcac
cagccccctc ctggcccctg tcatcttctg 541 tcccttccca gaaaacctac
cagggcagct acggtttccg tctgggcttc ttgcattctg 601 ggacagccaa
gtctgtgact tgcacgtact cccctgccct caacaagatg ttttgccaac 661
tggccaagac ctgccctgtg cagctgtggg ttgattccac acccccgccc ggcacccgcg
721 tccgcgccat ggccatctac aagcagtcac agcacatgac ggaggttgtg
aggcgctgcc 781 cccaccatga gcgctgctca gatagcgatg gtctggcccc
tcctcagcat cttatccgag 841 tggaaggaaa tttgcgtgtg gagtatttgg
atgacagaaa cacttttcga catagtgtgg 901 tggtgcccta tgagccgcct
gaggttggct ctgactgtac caccatccac tacaactaca 961 tgtgtaacag
ttcctgcatg ggcggcatga accggaggcc catcctcacc atcatcacac 1021
tggaagactc cagtggtaat ctactgggac ggaacagctt tgaggtgcgt gtttgtgcct
1081 gtcctgggag agaccggcgc acagaggaag agaatctccg caagaaaggg
gagcctcacc 1141 acgagctgcc cccagggagc actaagcgag cactgcccaa
caacaccagc tcctctcccc 1201 agccaaagaa gaaaccactg gatggagaat
atttcaccct tcagatccgt gggcgtgagc 1261 gcttcgagat gttccgagag
ctgaatgagg ccttggaact caaggatgcc caggctggga 1321 aggagccagg
ggggagcagg gctcactcca gccacctgaa gtccaaaaag ggtcagtcta
http://www.ncbi.nlm.nih.gov/entrez/viewer.fcgi?val=8400737&from=354&to=552&view=gbwithpartshttp://www.ncbi.nlm.nih.gov/genome/sts/sts.cgi?uid=154969http://www.ncbi.nlm.nih.gov/entrez/viewer.fcgi?val=8400737&from=368&to=626&view=gbwithpartshttp://www.ncbi.nlm.nih.gov/genome/sts/sts.cgi?uid=154963http://www.ncbi.nlm.nih.gov/entrez/viewer.fcgi?val=8400737&from=729&to=803&view=gbwithpartshttp://www.ncbi.nlm.nih.gov/genome/sts/sts.cgi?uid=273171http://www.ncbi.nlm.nih.gov/entrez/viewer.fcgi?val=8400737&from=1004&to=1202&view=gbwithpartshttp://www.ncbi.nlm.nih.gov/genome/sts/sts.cgi?uid=272633http://www.ncbi.nlm.nih.gov/entrez/viewer.fcgi?val=8400737&from=1008&to=1082&view=gbwithpartshttp://www.ncbi.nlm.nih.gov/genome/sts/sts.cgi?uid=155620http://www.ncbi.nlm.nih.gov/entrez/viewer.fcgi?val=8400737&from=1503&to=1619&view=gbwithpartshttp://www.ncbi.nlm.nih.gov/genome/sts/sts.cgi?uid=82485http://www.ncbi.nlm.nih.gov/entrez/viewer.fcgi?val=8400737&from=2503&to=2579&view=gbwithpartshttp://www.ncbi.nlm.nih.gov/genome/sts/sts.cgi?uid=59997http://www.ncbi.nlm.nih.gov/entrez/viewer.fcgi?val=8400737&from=2612&to=2617&view=gbwithpartshttp://www.ncbi.nlm.nih.gov/entrez/viewer.fcgi?val=8400737&itemID=3&view=gbwithparts
-
1381 cctcccgcca taaaaaactc atgttcaaga cagaagggcc tgactcagac
tgacattctc 1441 cacttcttgt tccccactga cagcctccca cccccatctc
tccctcccct gccattttgg 1501 gttttgggtc tttgaaccct tgcttgcaat
aggtgtgcgt cagaagcacc caggacttcc 1561 atttgctttg tcccggggct
ccactgaaca agttggcctg cactggtgtt ttgttgtggg 1621 gaggaggatg
gggagtagga cataccagct tagattttaa ggtttttact gtgagggatg 1681
tttgggagat gtaagaaatg ttcttgcagt taagggttag tttacaatca gccacattct
1741 aggtaggtag gggcccactt caccgtacta accagggaag ctgtccctca
tgttgaattt 1801 tctctaactt caaggcccat atctgtgaaa tgctggcatt
tgcacctacc tcacagagtg 1861 cattgtgagg gttaatgaaa taatgtacat
ctggccttga aaccaccttt tattacatgg 1921 ggtctaaaac ttgaccccct
tgagggtgcc tgttccctct ccctctccct gttggctggt 1981 gggttggtag
tttctacagt tgggcagctg gttaggtaga gggagttgtc aagtcttgct 2041
ggcccagcca aaccctgtct gacaacctct tggtcgacct tagtacctaa aaggaaatct
2101 caccccatcc cacaccctgg aggatttcat ctcttgtata tgatgatctg
gatccaccaa 2161 gacttgtttt atgctcaggg tcaatttctt ttttcttttt
tttttttttt tttctttttc 2221 tttgagactg ggtctcgctt tgttgcccag
gctggagtgg agtggcgtga tcttggctta 2281 ctgcagcctt tgcctccccg
gctcgagcag tcctgcctca gcctccggag tagctgggac 2341 cacaggttca
tgccaccatg gccagccaac ttttgcatgt tttgtagaga tggggtctca 2401
cagtgttgcc caggctggtc tcaaactcct gggctcaggc gatccacctg tctcagcctc
2461 ccagagtgct gggattacaa ttgtgagcca ccacgtggag ctggaagggt
caacatcttt 2521 tacattctgc aagcacatct gcattttcac cccacccttc
ccctccttct ccctttttat 2581 atcccatttt tatatcgatc tcttatttta
caataaaact ttgctgcca
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Question 6 LOCUS BC016497 1492 bp mRNA linear ROD 29-JUN-2004
DEFINITION Mus musculus eukaryotic translation initiation factor 2,
subunit 1 alpha, mRNA (cDNA clone MGC:25484 IMAGE:4501321),
complete cds. ACCESSION BC016497 VERSION BC016497.1 GI:16741331
KEYWORDS MGC. SOURCE Mus musculus (house mouse) ORGANISM Mus
musculus Eukaryota; Metazoa; Chordata; Craniata; Vertebrata;
Euteleostomi; Mammalia; Eutheria; Euarchontoglires; Glires;
Rodentia; Sciurognathi; Muroidea; Muridae; Murinae; Mus. REFERENCE
1 (bases 1 to 1492) AUTHORS Strausberg,R.L., Feingold,E.A.,
Grouse,L.H., Derge,J.G., Klausner,R.D., Collins,F.S., Wagner,L.,
Shenmen,C.M., Schuler,G.D., Altschul,S.F., Zeeberg,B., Buetow,K.H.,
Schaefer,C.F., Bhat,N.K., Hopkins,R.F., Jordan,H., Moore,T.,
Max,S.I., Wang,J., Hsieh,F., Diatchenko,L., Marusina,K.,
Farmer,A.A., Rubin,G.M., Hong,L., Stapleton,M., Soares,M.B.,
Bonaldo,M.F., Casavant,T.L., Scheetz,T.E., Brownstein,M.J.,
Usdin,T.B., Toshiyuki,S., Carninci,P., Prange,C., Raha,S.S.,
Loquellano,N.A., Peters,G.J., Abramson,R.D., Mullahy,S.J.,
Bosak,S.A., McEwan,P.J., McKernan,K.J., Malek,J.A., Gunaratne,P.H.,
Richards,S., Worley,K.C., Hale,S., Garcia,A.M., Gay,L.J.,
Hulyk,S.W., Villalon,D.K., Muzny,D.M., Sodergren,E.J., Lu,X.,
Gibbs,R.A., Fahey,J., Helton,E., Ketteman,M., Madan,A.,
Rodrigues,S., Sanchez,A., Whiting,M., Madan,A., Young,A.C.,
Shevchenko,Y., Bouffard,G.G., Blakesley,R.W., Touchman,J.W.,
Green,E.D., Dickson,M.C., Rodriguez,A.C., Grimwood,J., Schmutz,J.,
Myers,R.M., Butterfield,Y.S., Krzywinski,M.I., Skalska,U.,
Smailus,D.E., Schnerch,A., Schein,J.E., Jones,S.J. and Marra,M.A.
TITLE Generation and initial analysis of more than 15,000
full-length human and mouse cDNA sequences JOURNAL Proc. Natl.
Acad. Sci. U.S.A. 99 (26), 16899-16903 (2002) PUBMED
12477932REFERENCE 2 (bases 1 to 1492) AUTHORS Strausberg,R. TITLE
Direct Submission JOURNAL Submitted (31-OCT-2001) National
Institutes of Health, Mammalian Gene Collection (MGC), Cancer
Genomics Office, National Cancer Institute, 31 Center Drive, Room
11A03, Bethesda, MD 20892-2590, USA REMARK NIH-MGC Project URL:
http://mgc.nci.nih.govCOMMENT Contact: MGC help desk Email:
[email protected] Tissue Procurement: The Cepko Laboratory cDNA
Library Preparation: Life Technologies, Inc. cDNA Library Arrayed
by: The I.M.A.G.E. Consortium (LLNL) DNA Sequencing by: Baylor
College of Medicine Human Genome Sequencing Center Center code:
BCM-HGSC
http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?id=10090http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=pubmed&dopt=Abstract&query_hl=1&list_uids=12477932http://mgc.nci.nih.gov/
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Web site: http://www.hgsc.bcm.tmc.edu/cdna/ Contact:
[email protected] Gunaratne, P.H., Garcia, A.M., Lu, X., Hulyk, S.W.,
Loulseged, H., Kowis, C.R., Sneed, A.J., Martin, R.G., Muzny, D.M.,
Nanavati, A.N., Gibbs, R.A. Clone distribution: MGC clone
distribution information can be found through the I.M.A.G.E.
Consortium/LLNL at: http://image.llnl.gov Series: IRAK Plate: 31
Row: k Column: 7 This clone was selected for full length sequencing
because it passed the following selection criteria: matched mRNA
gi: 13385623. FEATURES Location/Qualifiers source 1..1492
/organism="Mus musculus" /mol_type="mRNA" /db_xref="taxon:10090"
/clone="MGC:25484 IMAGE:4501321" /tissue_type="Eye, retina, mouse
strain C57Bl\6" /clone_lib="NIH_MGC_94" /lab_host="DH10B"
/note="Vector: pCMV-SPORT6" gene 1..1492 /gene="Eif2s1"
/note="synonyms: 0910001O23Rik, eIF2alpha, 35kDa"
/db_xref="GeneID:13665" /db_xref="MGI:95299" CDS 115..1062
/gene="Eif2s1" /codon_start=1 /product="eukaryotic translation
initiation factor 2, subunit 1 alpha" /protein_id="AAH16497.1"
/db_xref="GI:16741332" /db_xref="GeneID:13665" /db_xref="MGI:95299"
/translation="MPGLSCRFYQHKFPEVEDVVMVNVRSIAEMGAYVSLLEYNNIEG
MILLSELSRRRIRSINKLIRIGRNECVVVIRVDKEKGYIDLSKRRVSPEEAIKCEDKF
TKSKTVYSILRHVAEVLEYTKDEQLESLFQRTAWVFDDKYKRPGYGAYDAFKHAVSDP
SILDSLDLNEDEREVLINNINRRLTPQAVKIRADIEVACYGYEGIDAVKEALRAGLNC
STETMPIKINLIAPPRYVMTTTTLERTEGLSVLNQAMAVIKEKIEEKRGVFNVQMEPK
VVTDTDETELARQLERLERENAEVDGDDDAEEMEAKAED" misc_difference 1468..1492
/gene="Eif2s1" /note="polyA tail: 25 bases do not align to the
mouse genome; Differences found between this sequence and the mouse
C57BL/6J genome (build 35) are described in misc_difference
features below" ORIGIN 1 gcgaagcgca cgctgaggag cttcggcgtc
cggggttggt gttccgagtc tcctcgttgc
http://www.hgsc.bcm.tmc.edu/cdna/http://image.llnl.gov/http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?id=10090http://www.ncbi.nlm.nih.gov/entrez/viewer.fcgi?val=16741331&itemID=2&view=gbwithpartshttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=gene&cmd=retrieve&dopt=graphics&list_uids=13665http://www.informatics.jax.org/searches/accession_report.cgi?id=MGI:95299http://www.ncbi.nlm.nih.gov/entrez/viewer.fcgi?val=16741331&itemID=1&view=gbwithpartshttp://www.ncbi.nlm.nih.gov/entrez/viewer.fcgi?val=AAH16497.1http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=gene&cmd=retrieve&dopt=graphics&list_uids=13665http://www.informatics.jax.org/searches/accession_report.cgi?id=MGI:95299http://www.ncbi.nlm.nih.gov/entrez/viewer.fcgi?val=16741331&itemID=4&view=gbwithparts
-
61 cactaagcag cacggcgtgg gaccctactt cgggattcac acatccactt
cagaatgccg 121 gggctaagtt gtagatttta tcaacacaaa tttcctgagg
tggaagatgt agtgatggtg 181 aatgtaagat ccattgctga aatgggggcc
tatgtcagct tgttggaata taataacatt 241 gaaggcatga ttcttcttag
tgaattatcc agacgacgta tccgttctat aaacaaactg 301 atccgaattg
gcagaaatga atgtgttgtt gtcattagag tggataaaga aaaaggatat 361
atagatttgt caaaaagaag agtttctcca gaggaagcaa tcaaatgtga ggacaaattc
421 acaaaatcca aaactgttta tagcattctt cgccatgttg ctgaggtatt
agaatatacc 481 aaggatgagc agctggagag cctgttccag aggactgcct
gggtcttcga tgacaagtac 541 aagagacctg gatacggtgc ctacgatgct
tttaagcatg cagtctcaga cccatctatc 601 ttggatagtt tagatttgaa
tgaagatgaa agagaagtac tcattaataa tatcaatagg 661 cgtttgaccc
cacaagcggt caaaattcga gcagatattg aagtagcttg ctatggttat 721
gaaggcattg atgctgtaaa agaagccctg agggcaggtt tgaattgttc tacagaaacc
781 atgcccatca agattaatct aatagctcca cccaggtatg tgatgacaac
aacgaccctg 841 gagaggacag aaggcctgtc tgtcctcaat caggctatgg
cagttatcaa agagaagatc 901 gaggagaaga ggggcgtctt caatgttcag
atggagccca aagtggtcac agatacagat 961 gagactgaac ttgcaaggca
gctggaacgg ctggagagag aaaatgcaga agtggatgga 1021 gatgatgatg
cagaagaaat ggaagccaaa gctgaagatt aacgttttgg caaacagtcc 1081
aatttaagga gtacgaagca gcccttcctg gctgtcaacc ctagacttga gagttttcca
1141 gtattgaaaa cttcaaagct gaatattttt atttccaagt atttaagtat
tcaacaagcc 1201 agaatctaaa tgccctcctt catgtcagct gtttccacat
agtggctcta acacctcaag 1261 catttttcaa gggagtggct tgatttgacc
agagacaaat cttaaacagc agtcctaata 1321 ttgggcctac agtttccatt
tctcatgtct ctggaatggc acccttatgg ttcagagatt 1381 taccagggac
tccaaacaca aacaatccca atctttctat ataaaatgta ttcaagcaaa 1441
catcaaataa atttctggga tatttaaaaa aaaaaaaaaa aaaaaaaaaa aa
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Question 6.
Name: eIF2a
Conformation: linear
Overhang: five_prime, three_prime, blunt
Minimum Site Length: 6 bases
Maximum Number of Cuts: 2
Included: all commercial, prototypes only
Noncutters: AarI, AatII, AccI, AflII, AflIII, AgeI, AlwNI, ApaI,
ApaLI, AscI, AsuII, AvaI, AvrII, BaeI, BalI, BamHI, BcgI, BclI,
BfiI, BglII, BsaAI, BsaBI, BsePI, BseSI, BseYI, BsgI, Bsp1407I,
BspHI, BspLU11I, BsrBI, BstEII, BstXI, BtrI, BtsI, CfrI, Cfr10I,
ClaI, DrdI, Eam1105I, EciI, Eco47III, EcoRI, EcoRV, Esp3I, FseI,
FspAI, HaeII, HindIII, HpaI, KpnI, MfeI, MluI, NaeI, NarI, NcoI,
NdeI, NheI, NotI, NruI, OliI, PI-PspI, PI-SceI, PacI, PfoI, PmaCI,
PmeI, PshAI, PsiI, PsrI, PstI, PvuI, RsrII, SacI, SacII, SalI,
SapI, SexAI, SfiI, SgfI, SgrAI, SmaI, SnaBI, SpeI, SrfI, Sse8387I,
SwaI, TaqII, Tth111I, XbaI, XhoI
Name Sequence Site Length Overhang Frequency Cut Positions
HindII GTYRAC 6 blunt 1 1116
ScaI AGTACT 6 blunt 1 638
StuI AGGCCT 6 blunt 1 854
XmnI GAANNNNTTC 6 blunt 1 381
AclI AACGTT 6 five_prime 1 1062
BspMI ACCTGC 6 five_prime 1 745
Eco31I GGTCTC 6 five_prime 1 537
EcoNI CCTNNNNNAGG 6 five_prime 1 868
PpuMI RGGWCCY 7 five_prime 1 80
SanDI GGGWCCC 7 five_prime 1 80
StyI CCWWGG 6 five_prime 1 479
XhoII RGATCY 6 five_prime 1 187
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Name Sequence Site Length Overhang Frequency Cut Positions
BglI GCCNNNNNGGC 6 three_prime 1 751
BsrDI GCAATG 6 three_prime 1 191
MmeI TCCRAC 6 three_prime 1 203
NspI RCATGY 6 three_prime 1 580
PflMI CCANNNNNTGG 6 three_prime 1 1241
SduI GDGCHC 6 three_prime 1 938
SphI GCATGC 6 three_prime 1 580
XcmI CCANNNNNNNNNTGG 6 three_prime 1 198
MslI CAYNNNNRTG 6 blunt 2 337, 1364
SspI AATATT 6 blunt 2 1164, 1319
AcyI GRCGYC 6 five_prime 2 26, 914
BbvCI CCTCAGC 7 five_prime 2 13, 461
Bpu10I CCTNAGC 6 five_prime 2 13, 461
DraII RGGNCCY 6 five_prime 2 80, 206
Ksp632I CTCTTC 6 five_prime 2 372, 901
SmlI CTYRAG 6 five_prime 2 1126, 1255
TatI WGTACW 6 five_prime 2 536, 636
VspI ATTAAT 6 five_prime 2 644, 794
AloI GAACNNNNNNTCC 7 three_prime 2 487, 519
BciVI GTATCC 6 three_prime 2 289, 544
BplI GAGNNNNNCTC 6 three_prime 2 623, 655
BsaXI ACNNNNNCTCC 6 three_prime 2 487, 517
BsmI GAATGC 6 three_prime 2 119, 443
DraIII CACNNNGTG 6 three_prime 2 76, 1241
Eco57I CTGAAG 6 three_prime 2 93, 1073
FalI AAGNNNNNCTT 6 three_prime 2 1087, 1119
Hin4I GAYNNNNNVTC 6 three_prime 2 263, 295
PpiI GAACNNNNNCTC 7 three_prime 2 487, 519
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Sites de restriction pour pOPRSV/MCS : Enzyme Nombre de
nucléotides additionnels Site de restriction requis en 5’ pour
couper cette séquence Cla I : 2 EcoR V : 1 Kpn I : 2 Not I : 8 Sma
I : 3 Spe I : 2 Xba I : 1 Xho I : 1