pAAV2-7M8 vector (Cat. No.: V012820)

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Basic Information

Note: AAV packaging plasmid, expressing Rep2 and 7M8 capsid. 7mer insertion in the AAV2 cap. It engineered variant of the AAV2 serotype, designed for highly efficient retinal transduction following intravitreal injection. Its key advantage is the ability to infect a broad range of retinal cells, including inner retinal neurons and photoreceptors, without the need for a subretinal delivery approach. In the provided study, it was the most effective vector for delivering the anti-VEGF transgene (KH902), demonstrating superior transduction and therapeutic efficacy in mouse models of neovascular eye disease.

Name:: pAAV2-7M8

Antibiotic Resistance:: Ampicillin

Length:: 8369 bp

Type:: Viral Expression & Packaging Vectors

Replication origin:: ori

Growth Strain(s):: DH10B

Growth Temperature:: 37℃

$ 198.2

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

Resources

Plasmid Protocol

1. Centrifuge at 5,000×g for 5 min.

2. Carefully open the tube and add sterile water to dissolve the DNA: add 20 μl for 5 μg plasmid, and 100 μl for 100 μg plasmid.

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

Cheng SY, Luo Y, Malachi A, Ko J, Su Q, Xie J, Tian B, Lin H, Ke X, Zheng Q, Tai PWL, Gao G, Punzo C. Low-Dose Recombinant Adeno-Associated Virus-Mediated Inhibition of Vascular Endothelial Growth Factor Can Treat Neovascular Pathologies Without Inducing Retinal Vasculitis. Hum Gene Ther. 2021 Jul;32(13-14):649-666. doi: 10.1089/hum.2021.132. PMID: 34182803; PMCID: PMC8312021.

pAAV2-7M8 vector (Cat. No.: V012820) Sequence

LOCUS       pAAV2-7M8               8369 bp    DNA     circular SYN 09-MAY-2026
DEFINITION  Exported.
ACCESSION   V012820
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8369)
  TITLE     Direct Submission
REFERENCE   2  (bases 1 to 8369)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..8369
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   8..110
                     /label=ITR
                     /note="inverted terminal repeat of human adenovirus
                     serotype 5"
     misc_feature    121..252
                     /label=P5 promoter
                     /note="/vntifkey=21"
     CDS             258..2114
                     /gene="Rep78"
                     /label=Protein Rep78
                     /note="Protein Rep78 from Adeno-associated virus 2 (isolate
                     Srivastava/1982). Accession#: Q89268"
     CDS             2134..4371
                     /codon_start=1
                     /label=AAV2.7m8
                     /note="AAV2.7m8 is an engineered AAV2 capsid variant 
                     created by in vivo directed evolution. It carries a 7‑amino
                     acid insertion (LALGETTRP) at position 588 and a V708I 
                     point mutation, which enable highly efficient outer retinal
                     transduction from a single intravitreal injection, 
                     bypassing the inner limiting membrane and penetrating deep 
                     into photoreceptors and RPE."
                     /translation="MAADGYLPDWLEDTLSEGIRQWWKLKPGPPPPKPAERHKDDSRGL
                     VLPGYKYLGPFNGLDKGEPVNEADAAALEHDKAYDRQLDSGDNPYLKYNHADAEFQERL
                     KEDTSFGGNLGRAVFQAKKRVLEPLGLVEEPVKTAPGKKRPVEHSPVEPDSSSGTGKAG
                     QQPARKRLNFGQTGDADSVPDPQPLGQPPAAPSGLGTNTMATGSGAPMADNNEGADGVG
                     NSSGNWHCDSTWMGDRVTTTSTRTWALPTYNNHLYKQISSQSGASNDNHYFGYSTPWGY
                     FDFNRFHCHFSPRDWQRLINNNWGFRPKRLNFKLFNIQVKEVTQNDGTTTIANNLTSTV
                     QVFTDSEYQLPYVLGSAHQGCLPPFPADVFMVPQYGYLTLNNGSQAVGRSSFYCLEYFP
                     SQMLRTGNNFTFSYTFEDVPFHSSYAHSQSLDRLMNPLIDQYLYYLSRTNTPSGTTTQS
                     RLQFSQAGASDIRDQSRNWLPGPCYRQQRVSKTSADNNNSEYSWTGATKYHLNGRDSLV
                     NPGPAMASHKDDEEKFFPQSGVLIFGKQGSEKTNVDIEKVMITDEEEIRTTNPVATEQY
                     GSVSTNLQRGNLALGETTRPARQAATADVNTQGVLPGMVWQDRDVYLQGPIWAKIPHTD
                     GHFHPSPLMGGFGLKHPPPQILIKNTPVPANPSTTFSAAKFASFITQYSTGQVSVEIEW
                     ELQKENSKRWNPEIQYTSNYNKSINVDFTVDTNGVYSEPRPIGTRYLTRNL"
     CDS             4213..4224
                     /label=WELQut site
                     /note="WELQut protease recognition and cleavage site"
     misc_feature    4467..4598
                     /label=P5  promoter
                     /label=P5 promoter
                     /note="/vntifkey=21"
     promoter        4562..4597
                     /label=Truncated P5
                     /label=Truncated\P5
                     /note="/vntifkey=29"
     rep_origin      5150..5663
                     /label=M13 ori
                     /note="M13 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
     promoter        6410..6514
                     /label=AmpR promoter
     CDS             join(6515..6583,6584..7375)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     rep_origin      7546..8134
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"