pRNATin-H1.2/Retro siRNA expression vector is compatible with Clontech Retro-X Expression System. It uses an inducible H1 promoter for siRNA expression. The promoter contains the tetracycline operator (TetO1). As a competitor, tetracycline or doxycycline can bind this operator to remove the blockade of tetracycline repressor (TetR) from H1 promoter and induce the transcription of siRNA.The vector also carries a neomycin resistance gene for establishing stable transfection cell line and a cGFP marker for tracking transfection efficiency, both genes are under the control of SV40 promoter. The vector uses CMV enhancer/promoter joined MSV 5’LTR (CMV/MSV 5LTR) and MSV 3LTR for viral transcription and packaging.
| 载体基本信息 | |||
| 载体名称 | pRNATin-H1.2/Retro | 载体抗性 | Ampicillin |
|---|---|---|---|
| 载体长度 | 7706 bp | 载体类型 | RNAi vectors |
| 筛选标记 | Neomycin | 启动子 | H1.2 |
| 克隆方法 | Enzyme digestion and ligation | 5引物 | CCAACTTGTTTATTGCAGCT |
| 3引物 | TAATACGACTCACTATAGGG | 载体标签 | cGFP |
pRNATin-H1.2/Retro 载体载体图谱
pRNATin-H1.2/Retro 载体载体序列
LOCUS Exported 7706 bp ds-DNA circular SYN 13-JAN-2022
DEFINITION synthetic circular DNA
ACCESSION .
VERSION .
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 7706)
TITLE Direct Submission
FEATURES Location/Qualifiers
source 1..7706
/organism="synthetic DNA construct"
/mol_type="other DNA"
promoter 309..512
/note="CMV promoter"
/note="human cytomegalovirus (CMV) immediate early
promoter"
LTR 513..688
/note="5 LTR (truncated)"
/note="truncated long terminal repeat from Moloney murine
sarcoma virus"
misc_feature 751..950
/note="MMLV Psi"
/note="packaging signal of Moloney murine leukemia virus
(MMLV)"
CDS 1151..1567
/note="gag (truncated)"
/note="truncated Moloney murine leukemia virus (MMLV) gag
gene lacking the start codon"
promoter 1604..1920
/note="SV40 promoter"
/note="SV40 enhancer and early promoter"
rep_origin 1771..1906
/note="SV40 ori"
/note="SV40 origin of replication"
misc_feature 2665..3238
/note="IRES"
/note="internal ribosome entry site (IRES) of the
encephalomyocarditis virus (EMCV)"
CDS 3270..4064
/codon_start=1
/gene="aph(3)-II (or nptII)"
/product="aminoglycoside phosphotransferase from Tn5"
/note="NeoR/KanR"
/note="confers resistance to neomycin, kanamycin, and G418
(Geneticin(R))"
/translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
polyA_signal 4238..4359
/note="SV40 poly(A) signal"
/note="SV40 polyadenylation signal"
protein_bind 4421..4439
/bound_moiety="tetracycline repressor TetR"
/note="tet operator"
/note="bacterial operator O1 for the tetR and tetA genes"
promoter complement(4515..4533)
/note="T7 promoter"
/note="promoter for bacteriophage T7 RNA polymerase"
promoter 5103..5432
/note="SV40 promoter"
/note="SV40 enhancer and early promoter"
rep_origin 5283..5418
/note="SV40 ori"
/note="SV40 origin of replication"
rep_origin complement(5725..6313)
/direction=LEFT
/note="ori"
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
CDS complement(6484..7344)
/codon_start=1
/gene="bla"
/product="beta-lactamase"
/note="AmpR"
/note="confers resistance to ampicillin, carbenicillin, and
related antibiotics"
/translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
LIKHW"
promoter complement(7345..7449)
/gene="bla"
/note="AmpR promoter"
enhancer 7635..308
/note="CMV enhancer"
/note="human cytomegalovirus immediate early enhancer"
