DNA transfection and adenovirus construction

(50 μg linearized DNA required)

Produces plaque isolated adenoviral clones (up to 12 clones).

$520

$2,985 (includes construction, amplification, large scale purification, and titer)

Plaque Amplification

Up to 12, 1 ml amplified clones of adenovirus (primary lysate).

$75

$125

Adenovirus Purification & Expansion

(large scale)

Yield varies greatly between constructs.  On average 2.5x10¹³ particles are typical. When amplifying lysate, please bring 50-100 μl primary lysate if possible. When amplifying purified virus, please bring 10 μl. (Note: for Ad constructs generated by Vector Core, we archive a small aliquot of primary lysate for further expansion, if requested.) When amplifying purified adenovirus that has been serially propagated it may be wise to isolate viral clones (with a plaque amplification), in order to avoid any possible problems with replication competent adenovirus.

$1,050

$2,005 (includes titer)

Standard vector glycerol stock

(10¹² particles, includes titer)

Each vial contains 0.25 mls of 4x10¹² particles/ml. A titer is included.

$130

$215

Plaque Assay (titer)

For titer of an adenovirus in PFU/ml (note: the same procedure is used for clonal isolation of possibly replication competent viruses)

$165

$275

Crude Virus (lysate, 50 ml)

Each prep is 50 mls of adenoviral lysate which has been clarified of cell debris, but not purified or concentrated.

$90

$155

Name

Map

Sequence

Description

Ad5 CMV-cre

NOTE: Available only to UM investigators. Cre recombinase is a Type I topoisomerase from P1 bacteriophage that catalyzes site-specific recombination of DNA between loxP sites. An E1/E3 deleted adenovirus type 5 vector expressing cre recombinase under CMV promoter control.

Ad5 HSV-tk

Herpes virus thymidine kinase (HSV-tk) is used in suicide gene therapy. It has found success in treating prostate cancer. Lyoma virus enhancer, HSV-tk enhancer and promoter, HSV thymidine kinase. E1 and partial E3 deleted.

Ad5 wt p53

p53 is critical for DNA repair and apoptosis. Mutations of p53 are associated with many forms of human malignancy. E1/E3 deleted, CMV promoter, expressing human p53 wild type.

Ad5 CMV hIL10

Interleukin (IL)-10 is a cytokine with potent anti-inflammatory properties. An E1/E3 deleted adenovirus type 5 vector expressing human interleukin 10 (hIL-10) under CMV promoter control.

Ad5 p16

p16 is a cyclin-dependent kinase inhibitor that has shown prognostic utility in some human cancers. Deletion or mutation of the p16 gene leads to tumor development or tumor progression. E1 and partial E3 deleted with CMV promoter.

Ad5 p21

p21 is a cyclin-dependent kinase inhibitor. It is a key effector of cellular senescence downstream of the tumor suppressor p53. E1 and partial E3 deleted with CMV promoter.

Ad5 p27

p27 is a cyclin-dependent kinase inhibitor with many putative functions in normal and neoplastic cells. It has a role in inhibiting cell proliferation, with prognostic and therapeutic implications for human cancer. E1 and partial E3 deleted with CMV promoter.

Ad5 RSV TGF beta

Transforming growth factor beta (TGF beta) is a multifunctional polypeptide which regulates normal cell growth, development, and tissue remodeling following injury. Abnormalities in TGF beta have been tied to tumorigenicity. E1 and partial E3 deleted, RSV promoter, expressing constituently active transforming growth factor beta.

Ad5 BclxS

Recombinant adenovirus also called Ad52bclxs/loxp. Bclx is an anti-apoptotic oncogene that is activated in tumor growth. E1 and partial E3 deleted adenoviral vector, RSV LTR promoter, and expresses human Bclx short (BclxS).

Name

Map

Sequence

Description

Ad5 CMV eGFP .dlE3

E1/E3 deleted adenoviral reporter vector, CMV promoter, expresses green fluorescent protein.

Ad5 CMV-Luc

An adenoviral reporter vector expressing luciferase (firefly). E1/E3 deleted with CMV promoter.

Ad5 CMV nt- β-gal .dlE3

Adenoviral reporter vector, E1/E3 deleted, CMV promoter, expresses nuclear targeted β-galactosidase (LacZ). Nuclear and cytoplasm localized β-gal may be detected in some cell types.

Ad5 RSV nt- β-gal .dlE3

N/A

N/A

Adenoviral reporter vector, E1/E3 deleted, RSV promoter, expresses nuclear targeted β-galactosidase (LacZ). Nuclear and cytoplasm localized β-gal may be detected in some cell types.

Ad5 CMVnt-LacZ

N/A

N/A

E1 and partial E3 deleted adenoviral reporter vector, CMV promoter, expresses nuclear targeted β-galactosidase (LacZ).

Ad5 RSV nt-LacZ

Adenoviral reporter vector, E1 and partial E3 deleted, RSV LTR promoter, expresses nuclear targeted β-galactosidase (LacZ).

Ad5 RSV LacZ

Adenoviral reporter vector, E1 and partial E3 deleted, RSV LTR promoter, expresses cytoplasm localized β-galactosidase (LacZ).

Name

Map

Sequence

Description

Ad5CMV.dlE3

Recombinant adenovirus, also called AdCMVpLpA(-)loxp.dlE3 is an E1/E3 deleted, empty control vector with a CMV promoter, SV40 polyA site.

Ad5RSV.dlE3

Recombinant adenovirus, also called AdRSVpLpA(-)loxp.dlE3 is an E1/E3 deleted, empty control vector with an RSV promoter, SV40 polyA site.

Ad5EF1alpha

This is an empty control vector. E1 and partial E3 deleted, EF1 alpha promoter, bovine growth hormone polyA site. The shuttle used to generate this virus was pAdEF1 alpha loxP.

Ad5CMV

Recombinant adenovirus, also called AdCMVpLpA(-)loxp is an E1 and partial E3 deleted empty control vector with an EF1 alpha promoter, bovine growth hormone polyA site and dl309 backbone.

Ad5 pL(+)loxP

This is an empty control vector. E1 and partial E3 deleted. Does not contain promoter or polyA site.

Name

Map

Sequence

Description

Ad5 wt

Standard, replication competent wild-type adenovirus (serotype 5).

Ad5 sub 360

Replication competent, partial E3 deleted adenovirus with dl309 backbone. Endogenous E1 region with MLP substituted for E1A promoter.

Ad5 dl 7001

The E3 region of the adenovirus genome encodes at least nine alternatively spliced RNAs and at least seven distinct proteins. In order to better characterize the function of the E3 proteins a mutant adenovirus devoid of all E3 coding sequences was constructed by Wold and colleagues and this deletion mutant was designated dl7001. This virus has a deletion spanning nt 362-3382 of the Ad5 E3 transcription unit. Despite the absence of E3 proteins, dl7001 replicates well in adenoviral packaging cell lines and has proven to be a useful genomic backbone for the derivation of recombinant Ad5 vectors.

Name

Map

Sequence

Description

pACCMV2

Adenoviral shuttle plasmid with CMV promoter, pUC19 polylinker, SV40 splice/polyA, loxP and unique restriction tag SwaI, PmeI, NheI for linearization. Updated version of pACCMVpLpA(-)loxp-SSP. Shuttle used to produce Ad5 E1/E3-deleted virus.

pACRSV2

Adenoviral shuttle plasmid with RSV promoter, pUC19 polylinker, SV40 splice/polyA, loxP and unique restriction tag SwaI, PmeI, NheI for linearization. Updated version of pARSVpLpA(-)loxp-SSP. Shuttle used to produce Ad5 E1/E3-deleted virus.

pAdEF1 alpha loxP

Adenoviral shuttle plasmid with EF1 alpha promoter, polylinker, bovine growth hormone polyA site, loxP and unique restriction site NheI for linearization. Shuttle used to produce Ad5 E1/E3-deleted virus.

pACpL+loxP-SSP

Adenoviral shuttle plasmid with pUC19 polylinker and unique restriction tag SwaI, PmeI, NheI for linearization. Does not contain promoter or polyA site. Shuttle used to produce Ad5 E1/E3-deleted virus.

pACCMVpLpA(-) loxP-SSP

Adenoviral shuttle plasmid with CMV promoter, pUC19 polylinker, SV40 splice/polyA, loxP and unique restriction tag SwaI, PmeI, NheI for linearization. Shuttle used to produce Ad5 E1/E3-deleted virus.

pACRSVpLpA(-) loxP-SSP

Adenoviral shuttle plasmid with RSV promoter, pUC19 polylinker, SV40 splice/polyA, loxP and unique restriction tag SwaI, PmeI, NheI for linearization. Shuttle used to produce Ad5 E1/E3-deleted virus.

pACCMVpLmP1 (-)loxP-SSP

Adenoviral shuttle plasmid with CMV promoter, pUC19 polylinker, mP1 splicing signal/poly A, loxP with unique restriction tag SwaI, SfiI, PmeI for linearization. Shuttle used to produce Ad5 E1/E3-deleted virus.