pCC1FOS vector (V008674)

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Price Information

Cat No.	Plasmid Name	Availability	Add to cart
V008674	pCC1FOS	In stock, instant shipping	Buy one, get one free!

Two tubes of lyophilized plasmid will be delivered, each tube is about 5µg.

Basic Vector Information

The pCC1FOS is a single copy vector that is inducible with a CopyControl Induction Solution (CopyControl Fosmid clones grown in TransforMax EPI300 cells can be amplified to 10-50 copies per cell). An EPI400/EPI300 strain is recommended. The protocol is as below:
a) Add 4 ml LB media into each test tube. Inoculate each tube with bacterial culture with antibiotic at the proper concentration.
b) Incubate the tubes at 37°C, shaking overnight.
c) Dilute the starting culture (from step b) 1:10 into antibiotic-supplemented fresh media.
d) Supplement induction solution (0.04% Arabinose), and grow the culture at 37°C for 4 h with vigorous shaking (approx. 250 rpm).
e) Isolate DNA from the induced culture cells as per the protocol provided.

Vector Name:: pCC1FOS

Antibiotic Resistance:: Chloramphenicol

Length:: 8139 bp

Type:: Cloning vector

Replication origin:: ori2

Source/Author:: EPICENTRE Biotechnologies.

Copy Number:: Single copy

Growth Strain(s):: EPI300/400

Growth Temperature:: 37℃

pCC1FOS vector Map

Copy Sequence View Map

Plasmid Protocol

1. Centrifuge at 5,000×g for 5 min.

2. Carefully open the tube and add 20 μl of sterile water to dissolve the DNA.

3. Close the tube and incubate for 10 minutes at room temperature.

4. Briefly vortex the tube and then do a quick spin to concentrate the liquid at the bottom. Speed is less than 5000×g.

5. Store the plasmid at -20 ℃.

6. The concentration of plasmid re-measurement sometimes differs from the nominal value, which may be due to the position of the lyophilized plasmid in the tube, the efficiency of the re-dissolution, the measurement bias, and adsorption on the wall of the tube, therefore, it is recommended to transform and extract the plasmid before using it

General Plasmid Transform Protocol

1. Take one 100μl of the competent cells and thaw it on ice for 10min, add 2μl of plasmid, then ice bath for 30min, then heat-shock it at 42℃ for 60s, do not stir, and then ice bath for 2min.

2. Add 900μl of LB liquid medium without antibiotics, and incubate at 37℃ for 45min (30℃ for 1-1.5 hours) with 180rpm shaking.

3. Centrifuge at 6000rpm for 5min, leave only 100μl of supernatant to resuspend the bacterial precipitate and spread it onto the target plasmid-resistant LB plate.

4. Invert the plate and incubate at 37℃ for 14h, or at 30℃ for 20h.

5. Pick a single colony into LB liquid medium, add the corresponding antibiotics, incubate at 220rpm for 14h, and extract the plasmid according to the experimental needs and the instructions of the plasmid extraction kit.

References

Lam KN, Cheng J, Engel K, Neufeld JD, Charles TC. Current and future resources for functional metagenomics. Front Microbiol. 2015 Oct 29;6:1196. doi: 10.3389/fmicb.2015.01196.

pCC1FOS vector Sequence

LOCUS       40924_9246        8139 bp DNA     circular SYN 17-DEC-2018
DEFINITION  Cloning vector pCC1FOS, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8139)
  AUTHORS   EPICENTRE Biotechnologies.
  TITLE     Direct Submission
  JOURNAL   Submitted (23-AUG-2007) 726 Post Road, Madison, WI 53713, USA
REFERENCE   2  (bases 1 to 8139)
  TITLE     Direct Submission
REFERENCE   3  (bases 1 to 8139)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Submitted 
            (23-AUG-2007) 726 Post Road, Madison, WI 53713, USA"
COMMENT     SGRef: number: 2; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..8139
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     288..304
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        311..329
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(443..459)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    complement(467..483)
                     /label=lac operator
                     /note="The lac repressor binds to the lac operator to
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(491..521)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    complement(536..557)
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     CDS             complement(780..1436)
                     /codon_start=1
                     /label=CmR
                     /note="chloramphenicol acetyltransferase"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     promoter        complement(1437..1539)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding
                     chloramphenicol acetyltransferase"
     rep_origin      2397..3011
                     /label=oriV
                     /note="origin of replication for the bacterial F plasmid"
     rep_origin      3087..3306
                     /label=ori2
                     /note="secondary origin of replication for the bacterial F 
                     plasmid; also known as oriS"
     CDS             3397..4149
                     /codon_start=1
                     /label=repE
                     /note="replication initiation protein for the bacterial F 
                     plasmid"
                     /translation="MAETAVINHKKRKNSPRIVQSNDLTEAAYSLSRDQKRMLYLFVDQ
                     IRKSDGTLQEHDGICEIHVAKYAEIFGLTSAEASKDIRQALKSFAGKEVVFYRPEEDAG
                     DEKGYESFPWFIKRAHSPSRGLYSVHINPYLIPFFIGLQNRFTQFRLSETKEITNPYAM
                     RLYESLCQYRKPDGSGIVSLKIDWIIERYQLPQSYQRMPDFRRRFLQVCVNEINSRTPM
                     RLSYIEKKKGRQTTHIVFSFRDITSMTTG"
     misc_feature    4155..4405
                     /label=incC
                     /note="incompatibility region of the bacterial F plasmid"
     CDS             4731..5903
                     /codon_start=1
                     /label=sopA
                     /note="partitioning protein for the bacterial F plasmid"
                     /translation="MFRMKLMETLNQCINAGHEMTKAIAIAQFNDDSPEARKITRRWRI
                     GEAADLVGVSSQAIRDAEKAGRLPHPDMEIRGRVEQRVGYTIEQINHMRDVFGTRLRRA
                     EDVFPPVIGVAAHKGGVYKTSVSVHLAQDLALKGLRVLLVEGNDPQGTASMYHGWVPDL
                     HIHAEDTLLPFYLGEKDDVTYAIKPTCWPGLDIIPSCLALHRIETELMGKFDEGKLPTD
                     PHLMLRLAIETVAHDYDVIVIDSAPNLGIGTINVVCAADVLIVPTPAELFDYTSALQFF
                     DMLRDLLKNVDLKGFEPDVRILLTKYSNSNGSQSPWMEEQIRDAWGSMVLKNVVRETDE
                     VGKGQIRMRTVFEQAIDQRSSTGAWRNALSIWEPVCNEIFDRLIKPRWEIR"
     CDS             5906..6874
                     /codon_start=1
                     /label=sopB
                     /note="partitioning protein for the bacterial F plasmid"
                     /translation="MKRAPVIPKHTLNTQPVEDTSLSTPAAPMVDSLIARVGVMARGNA
                     ITLPVCGRDVKFTLEVLRGDSVEKTSRVWSGNERDQELLTEDALDDLIPSFLLTGQQTP
                     AFGRRVSGVIEIADGSRRRKAAALTESDYRVLVGELDDEQMAALSRLGNDYRPTSAYER
                     GQRYASRLQNEFAGNISALADAENISRKIITRCINTAKLPKSVVALFSHPGELSARSGD
                     ALQKAFTDKEELLKQQASNLHEQKKAGVIFEAEEVITLLTSVLKTSSASRTSLSSRHQF
                     APGATVLYKGDKMVLNLDRSRVPTECIEKIEAILKELEKPAP"
     misc_feature    6950..7423
                     /label=sopC
                     /note="centromere-like partitioning region of the bacterial
                     F plasmid"
     misc_feature    complement(7683..8081)
                     /label=cos
                     /note="lambda cos site; allows packaging into phage lambda 
                     particles"
     protein_bind    complement(8099..8132)
                     /label=loxP
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."