我们所展示的质粒图谱主要是从文献和开放数据库中收集而来，主要是为了方便研究工作，其中一小部分质粒进行了质量控制，可供科学家使用。
所有的产品都严格仅供科学研究使用，不能应用于临床试验，包括人体摄入、注射或外用的药理学用途。
确保质粒的关键元件正确，但是我们并不能保证实验结果。页面展示的图谱序列为理论序列，可能与测序结果不一致，请自行比对后确定是否满足要求。(如果测过序，本页面一般会提供下载)
开放数据库中的大多数载体序列都没有被完全测序。如果实际序列与参考序列的相似度超过99%，则将其视为正确。
由于科学研究是在探索未知，具有很大的不确定性，在任何情况下，我们都不承担超出质粒本身的额外经济损失或责任。
- 载体名称:
- pSIREN_U6-shRNA_FF3-CMV-iRFP
- 载体抗性:
- Ampicillin
- 载体长度:
- 6896 bp
- 载体类型:
- Cloning vector
- 复制子:
- ori
- 载体来源:
- Li Y, Jiang Y, Chen H, Liao W, Li Z, Weiss R, Xie Z.
- 启动子:
- sacB
pSIREN_U6-shRNA_FF3-CMV-iRFP 载体图谱
质粒操作方法
1. 发货形式：质粒干粉(常温运输，存于-20度，请务必先转化提质粒后使用)
2. 收到质粒干粉后请先5000rpm离心1min，再加入20μl ddH2O溶解质粒;（质粒复测的浓度有时候与标称值差距较大，这可能是因为冻干质粒在管中的位置、复溶效率、测量偏差以及管壁的吸附导致，因此建议先转化提质粒后再使用）
3. 取1支100μl 感受态于冰上解冻10min，加入2μl质粒，再冰浴30min后，42℃热激60s，不要搅动，再冰浴2min；
4. 加入900μl无抗的LB液体培养基，180rpm震荡37℃培养45min (30℃培养1-1.5小时);
5. 6000rpm离心5min，仅留100μl上清液重悬细菌沉淀，并涂布至目标质粒抗性的LB平板上;
6. 将平板倒置37℃培养14h，如果要求是30℃则培养20h; (菌落过多则将质粒稀释后再转化；没有菌落则加入10μl质粒转化；建议不要直接转表达感受态， 要先转克隆感受态，重提质粒后再导入表达感受态);
7. 挑取单菌落至LB液体培养基中，加入对应抗生素，220rpm震荡培养14h，根据实验需要和质粒提取试剂盒说明书提取质粒。
pSIREN_U6-shRNA_FF3-CMV-iRFP 载体序列
LOCUS 40924_40452 6896 bp DNA circular SYN 18-DEC-2018
DEFINITION Cloning vector pSIREN_U6-shRNA_FF3-CMV-iRFP, complete sequence.
ACCESSION .
VERSION .
KEYWORDS .
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 6896)
AUTHORS Li Y, Jiang Y, Chen H, Liao W, Li Z, Weiss R, Xie Z.
TITLE Modular construction of mammalian gene circuits using TALE
transcriptional repressors
JOURNAL Unpublished
REFERENCE 2 (bases 1 to 6896)
AUTHORS Li Y, Jiang Y, Chen H, Liao W.
TITLE Direct Submission
JOURNAL Submitted (03-SEP-2014) Bioinformatics Division/Center for Synthetic
REFERENCE 3 (bases 1 to 6896)
TITLE Direct Submission
REFERENCE 4 (bases 1 to 6896)
AUTHORS .
TITLE Direct Submission
COMMENT SGRef: number: 1; type: "Journal Article"; journalName:
"Unpublished"
COMMENT SGRef: number: 2; type: "Journal Article"; journalName: "Submitted
(03-SEP-2014) Bioinformatics Division/Center for Synthetic "
COMMENT SGRef: number: 3; type: "Journal Article"
COMMENT ##Assembly-Data-START##
Sequencing Technology :: Sanger dideoxy sequencing
##Assembly-Data-END##
FEATURES Location/Qualifiers
source 1..6896
/mol_type="other DNA"
/organism="synthetic DNA construct"
primer_bind 78..94
/label=M13 fwd
/note="common sequencing primer, one of multiple similar
variants"
promoter 102..120
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
protein_bind 186..219
/label=loxP
/bound_moiety="Cre recombinase"
/note="Cre-mediated recombination occurs in the 8-bp core
sequence (GCATACAT)."
misc_feature 229..477
/label=Homo sapiens U6 promoter
/note="Homo sapiens U6 promoter"
misc_feature 484..536
/label=shRNA-FF3
/note="shRNA-FF3"
terminator complement(635..729)
/label=lambda t0 terminator
/note="transcription terminator from phage lambda"
enhancer 801..1104
/label=CMV enhancer
/note="human cytomegalovirus immediate early enhancer"
promoter 1105..1308
/label=CMV promoter
/note="human cytomegalovirus (CMV) immediate early
promoter"
CDS 1361..2308
/codon_start=1
/label=iRFP713
/note="phytochrome-based near-infrared fluorescent protein,
originally termed iRFP (Filonov et al., 2011)"
/translation="MAEGSVARQPDLLTCDDEPIHIPGAIQPHGLLLALAADMTIVAGS
DNLPELTGLAIGALIGRSAADVFDSETHNRLTIALAEPGAAVGAPITVGFTMRKDAGFI
GSWHRHDQLIFLELEPPQRDVAEPQAFFRRTNSAIRRLQAAETLESACAAAAQEVRKIT
GFDRVMIYRFASDFSGEVIAEDRCAEVESKLGLHYPASTVPAQARRLYTINPVRIIPDI
NYRPVPVTPDLNPVTGRPIDLSFAILRSVSPVHLEFMRNIGMHGTMSISILRGERLWGL
IVCHHRTPYYVDLDGRQACELVAQVLAWQIGVMEE"
polyA_signal 2326..2446
/label=SV40 poly(A) signal
/note="SV40 polyadenylation signal"
CDS complement(2465..3121)
/codon_start=1
/label=CmR
/note="chloramphenicol acetyltransferase"
/translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
protein_bind complement(3150..3183)
/label=loxP
/note="Cre-mediated recombination occurs in the 8-bp core
sequence (ATGTATGC) (Shaw et al., 2021)."
promoter 3198..3643
/label=sacB promoter
/note="sacB promoter and control region"
CDS 3644..5062
/codon_start=1
/label=SacB
/note="secreted levansucrase that renders bacterial growth
sensitive to sucrose"
/translation="MNIKKFAKQATVLTFTTALLAGGATQAFAKETNQKPYKETYGISH
ITRHDMLQIPEQQKNEKYQVPEFDSSTIKNISSAKGLDVWDSWPLQNADGTVANYHGYH
IVFALAGDPKNADDTSIYMFYQKVGETSIDSWKNAGRVFKDSDKFDANDSILKDQTQEW
SGSATFTSDGKIRLFYTDFSGKHYGKQTLTTAQVNVSASDSSLNINGVEDYKSIFDGDG
KTYQNVQQFIDEGNYSSGDNHTLRDPHYVEDKGHKYLVFEANTGTEDGYQGEESLFNKA
YYGKSTSFFRQESQKLLQSDKKRTAELANGALGMIELNDDYTLKKVMKPLIASNTVTDE
IERANVFKMNGKWYLFTDSRGSKMTIDGITSNDIYMLGYVSNSLTGPYKPLNKTGLVLK
MDLDPNDVTFTYSHFAVPQAKGNNVVITSYMTNRGFYADKQSTFAPSFLLNIKGKKTSV
VKDSILEQGQLTVNK"
promoter 5133..5237
/label=AmpR promoter
CDS 5238..6095
/codon_start=1
/label=AmpR
/note="beta-lactamase"
/translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
LIKHW"
rep_origin 6269..6857
/direction=RIGHT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"