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PubMedDate not supplied

Playing with fire: images of fire on toy packaging.

Despite prevention efforts designed to teach children that fire is not a toy, each year hundreds of children, primarily male, are injured while playing with fire. Mass-produced toys and games shape children's behaviors, and the fire images on toy packaging may send the message that fire is fun rather than dangerous. The purpose of this study was to determine the frequency of male fire-related injuries at our burn center and to investigate the use of fire images in toy packaging. An aisle-by-aisle search was conducted at a national toy store. Toys with fire images on their packaging were identified. Target gender and the setting in which the fire was displayed also were recorded. Only toys with clearly visible flames on their packaging were included. We reviewed our TRACS database from April 1997 to May 1999 for fire-related injuries to children admitted to our institution as a result of playing with fire. Review of toy stores revealed 404 toys with packaging containing fire images. Of these, 97% (393 of 404) were targeted to males. Video games were the leading category of flame-related packaging (208 of 404), followed by toy car/truck displays (84 of 404). Packaging for girls' toys had safe and contained settings, whereas boys' toys used settings that were uncontrolled and associated with speed and weapons. A total of 59 children, 52 males and 7 females, with a mean age of 10 +/- 0.6 years and mean TBSA burn of 12.3 +/- 2.1% were admitted to our facility with fire-related injuries during the study interval. Children imitate life with toys and use play to experiment with new behavior. Boys are receiving a powerful, consistent message from images of fire on toy packaging. These advertisements may help to contribute to the higher incidence of fire-related injuries in boys.

Accident Prevention↗
PubMed2005

Microbiological, biochemical and sensory assessment of mussels (Mytilus galloprovincialis) stored under modified atmosphere packaging.

AIMS: To determine the microbiological, biochemical and sensory changes of mussels during storage under aerobic, vacuum packaging (VP) and modified atmosphere packaging (MAP) conditions at 4 degrees C, and to determine shelf-life of mussels under the same packaging conditions using the above assessment parameters. METHODS AND RESULTS: Aqua-cultured mussels (Mytilus galloprovincialis) were obtained from a local culture farm, packaged aerobically under VP and MAP (50%/50% CO2/N2: M1, 80%/20% CO2/N2: M2, 40%/30%/30% CO2/N2/O2: M3), and stored at 4 degrees C. Quality evaluation was carried out using microbiological, chemical and sensory analyses. Microbiological results revealed that the M2 and VP delayed microbial growth compared with that of air-packaged samples. The effect was more pronounced for total viable count (TVC), Pseudomonas spp., lactic acid bacteria (LAB) and H2S-producing bacteria. TVC was reduced by 0.9-1.0, Pseudomonas spp. by 0.7-0.8, LAB by 1.0-2.2, H2S-producing bacteria by 0.7-1.2. Enterobacteriaceae were not significantly affected by MAP conditions. Of the chemical indices determined, the total volatile basic nitrogen and trimethylamine nitrogen values remained lower than the proposed acceptability limits of 35 mg N 100 g(-1) and 12 mg N 100 g(-1), respectively, after 15 days of storage. Both the VP and air-packaged mussel samples exceeded these limits. The thiobarbituric acid value of all MAP and VP mussels remained lower than the proposed acceptability limit of 1 mg malondialdehyde kg(-1). The air-packaged samples exceeded this limit. All samples retained desirable sensory characteristics during the first 8 days of storage. CONCLUSIONS: Based on odour and taste evaluation, the M1 and M3 samples remained acceptable until ca day 11-12, the M2 samples remained acceptable until ca day 14-15 days while the VP and air-packaged mussel samples remained acceptable until ca days 10-11 and 8-9 of storage respectively. Based primarily on sensory, but also on biochemical and microbiological parameters determined, M2 gas mixture was the most effective for mussel preservation achieving a shelf-life of ca 14-15 days. SIGNIFICANCE AND IMPACT OF THE STUDY: MAP (M2) can be used to increase the shelf-life of refrigerated mussels. A shelf-life extension of refrigerated mussels by ca 5-6 days under MAP may be obtained.

Animals↗
PubMed1993

Medication compliance in elderly outpatients using twice-daily dosing and unit-of-use packaging.

OBJECTIVE: To determine the effect of unit-of-use drug packaging of medications on compliance among elderly outpatients treated with complex medication regimens. DESIGN: Nonblind, randomized, clinical trial. SETTING: Geriatric outreach health centers in urban public housing units for independent-living elderly people. PATIENTS: Thirty-one patients (aged > or = 60 y), each taking three or more prescribed medications. Patients were randomly assigned to one of three study groups: group 1 (n = 12), no change in dosing or packaging; group 2 (n = 10), conventional packaging with twice-daily dosing; group 3 (n = 9), unit-of-use packaging with twice-daily dosing. INTERVENTION: A unit-of-use package consisting of a two-ounce plastic cup with a snap-on lid containing all medications to be taken at the time of dosing. MAIN OUTCOME MEASURES: Medication compliance was assessed monthly for six months using tablet counts. RESULTS: Medication compliance was significantly better in group 3 (92.6 percent) using unit-of-use packaging compared with either group 1 (79 percent) or group 2 (82.6 percent) (p = 0.017). Compliance did not differ between groups 1 and 2. CONCLUSIONS: In this small study of elderly outpatients taking three or more medications, unit-of-use packaging and twice-daily dosing improved medication compliance compared with conventional packaging.

Activities of Daily Living↗
PubMed2005

Obstruction of critical information on over-the-counter medication packages by external tags.

BACKGROUND: Over-the-counter (OTC) medication packages are important sources of information for consumers during product selection and use. Consumers may not be able to access information from OTC packages if external tags, namely price or anti-theft tags, are improperly placed. OBJECTIVE: To determine the amount and type of information concealed by anti-theft tags and price tags affixed on OTC drug packages. METHODS: A field study was performed by evaluating packages containing acetaminophen and combinations of acetaminophen in stores located in Houston. Five packages for 4 products selected from each store after an initial survey for presence of an external tag were examined. A data collection sheet was prepared that extracted the type and amount of information concealed by these tags. Data were analyzed by performing descriptive analyses to provide an understanding of the information obscured. RESULTS: A total of 24 stores were considered in the study, and 67 products and 285 packages were evaluated. External tags, both anti-theft and price tags, obscured significant amounts of information on the principal display panel (53.4%) and the Drug Facts panel (47.7%) of OTC packages. These tags concealed crucial information on various aspects of labels such as brand names (42.3%), product description (36.3%), warnings (51.5%), uses (10.4%), and purposes (7.2%). CONCLUSIONS: Results suggest that improper use of external tags clearly obscured important information on OTC medication packages necessary for consumers to make informed decisions regarding product selection and use. Tagging practices should be altered to allow consumers full access to drug information on the product.

Advertising↗
PubMed1999

Effect of irradiation, packaging, and postirradiation cooking on the thiamin content of chicken meat.

The effect of irradiation with X rays or electrons, irradiation and storage temperature, and postirradiation cooking on the thiamin content of vacuum- or air-packaged minced chicken meat was examined. Samples irradiated with 3-kGy X rays (50 Gy/min) or electrons (5 kGy/min) contained less thiamin than the control specimens, but no differences between both irradiation methods were detected. The thiamin content in samples stored and/or irradiated at 5 degrees C was between 13 and 24 microg per 100-g product lower than in samples stored and/or ionized at -18 degrees C. The same difference in thiamin content was found for specimens packaged in a vacuum or air package, respectively. Vacuum packaging lead to a greater loss of drip than air-packaged samples. The biggest loss of thiamin, 31.1 and 28.0% for X rays and electron beams, respectively, was measured for vacuum-packaged specimens stored and irradiated at 5 degrees C. Compared with the cooked minced chicken breast meat, a higher thiamin content (6 to 17 microg of thiamin per 100-g product) was obtained for the raw samples. When irradiation and vacuum packaging were compared as two separate preservation techniques, the two methods had approximately the same effect on the thiamin content of the minced chicken meat. The mean temperature of the samples after cooking was 87.2 +/- 4.9 degrees C. However, significant differences in internal temperature after cooking of the samples were measured between air- and vacuum-packaged samples.

Animals↗
PubMed1999

Centralized packaging of retail meat cuts: a review.

Centralized packaging of retail meat cuts is growing more popular because of its economies and potential to maintain quality, enhance safety, and extend the shelf life of fresh meat. Requirements for optimizing shelf life of centrally prepared retail cuts for periods up to 15 weeks are slightly different from those needed to extend the shelf life of fresh, chilled meat. Chilled meat primarily deteriorate at the cut or uncut muscle surface. In long-term storage, primal cuts are placed in an atmosphere saturated with carbon dioxide and containing very low residual oxygen. These cuts are held at -1.5+/-0.5 degrees C. When the meat is removed, it is fabricated into retail or food service cuts. New fresh surfaces are created in the process, revitalizing the meat's appearance. After being prepared for retail display, the meat normally has four more days of shelf life. Depending on the meat species, shelf life is usually limited by development of undesirable organoleptic changes, usually defects in color, which are independent of microbial presence. The microbes consist of a lactic acid bacterial population that maximizes under storage conditions at about 10(8) CFU/cm2 well before shelf life ends. Circumstances are different with centralized distribution of retail-ready fresh meat. The wholesale storage period following initial packaging of the retail cuts is about 20 to 30 days. Prepared products must withstand retail display for up to 2 days without further manipulation of package contents. Retail packages are simply moved from their storage container (usually a unit or overwrap containing a modified atmosphere) to retail display, where desirable meat color develops upon exposure to air. Three gas atmospheres have some potential to satisfy storage needs for centralized distribution of retail-ready packages: 100% CO2, 100% N2, or 70% N2 + 30% CO2. Shelf life is limited by undesirable changes in surfaces exposed at initial packaging, caused by growth of psychrotrophic bacteria. If 100% CO2 is used, these are all lactic acid bacteria (LAB). Therefore, initial bacterial numbers on the meat and storage temperature become critical to success. The most attractive storage option is 100% CO2 used at - 1.5 +/-0.5 degrees C. This review presents the reason for that recommendation, along with basic concepts of meat chemistry, a discussion of modified atmosphere packaging, meat microbiology, and current results with simulated centralized packaging of retail-ready meats.

Food Packaging↗
PubMed2000

Review of centralized packaging systems for distribution of retail-ready meat.

There is growing interest in centralized preparation of retail-ready meat cuts for distribution to widely dispersed retail stores due to the convenience of having high-quality ready-to-go products that are consistently provided to consumers at lower cost. Various centralized packaging techniques are described. Of all packaging techniques, master packaging is the most economical and shows promise for commercial application. Nevertheless, the master-packaging technique must be integrated with strict temperature control in a narrow range just above freezing (- 1.5 +/- 0.5 degrees C), good processing hygiene, and maintenance of a completely anoxic atmosphere in the package headspace throughout the distribution period to maximize storage life. Packaging using the CAPTECH process reduces the residual O2 present in the headspace to 300 ppm. Oxygen scavengers must be incorporated in the package to absorb the residual O2 and preserve the metmyoglobin reducing activity of meat tissues. Integration of all these technologies can provide a storage life of retail-ready meat up to 10 weeks in the master package followed by 3 days of retail display life. This extension of storage life is sufficient for transporting meat to distant markets.

Animals↗
PubMed2002

Effect of nitrogen gas packaging on the quality and microbial growth of fresh-cut vegetables under low temperatures.

Nitrogen (N2) gas packaging for fresh-cut vegetables (lettuce and cabbage) has been examined as a means of modified atmosphere packaging (MAP) for extending the shelf life of cut vegetables. Gas composition in enclosed packages that contained cut vegetables and were filled with 100% N2 had an oxygen (O2) concentration of 1.2 to 5.0% and a carbon dioxide (CO2) concentration of 0.5 to 3.5% after 5 days of storage. An atmosphere of low concentrations of O2 and high CO2 conditions occurred naturally in the package filled with N2 gas. Degradation of cut vegetables in terms of appearance was delayed by N2 gas packaging. Because of this effect, the appearance of fresh-cut vegetables packaged with N2 gas remained acceptable at temperatures below 5 degrees C after 5 days. Treatment with acidic electrolyzed water (AcEW) contributed to the acceptability of the vegetables' appearance at 5 and 10 degrees C in the air-packaging system. N2 gas packaging did not significantly affect the growth of microbial populations (total aerobic bacteria, coliform bacteria, Bacillus cereus, and psychrotrophic bacteria) in or on cut vegetables at 1, 5, and 10 degrees C for 5 days. Microbial growth in or on the cut vegetables was inhibited at 1 degrees C for 5 days regardless of atmospheric conditions.

Bacteria↗
PubMed2005

Shelf-life extension of refrigerated Mediterranean mullet (Mullus surmuletus) using modified atmosphere packaging.

The present work evaluated the quality and freshness characteristics and the effect of modified atmosphere packaging (MAP) on the shelf-life extension of refrigerated Mediterranean mullet using microbiological, biochemical, and sensory analyses. Fresh open sea red mullet (Mullus surmuletus) were packaged in four different atmospheres: M1, 10%/20%/70% (O2/ CO2/N2); M2, 10%/40%/50% (O2/CO2/N2); M3, 10%/60%/30% (O2/CO2/N2); identical fish samples were packaged in air. All fish were kept under refrigeration (4 +/- 0.5 degrees C) for 14 days. Of the three gas atmospheres, the 10%/40%/50% (M2) and 10%/ 60%/30% (M3) gas mixtures were the most effective for inhibiting growth of aerobic microflora in mullet samples until day 10 of refrigerated storage. H2S-producing bacteria and pseudomonads were part of the mullet microflora and their growth was partly inhibited under MAP conditions. Between these two bacterial groups, H2S-producing bacteria (including Shewanella putrefaciens) were dominant toward the end of the storage period, regardless of the packaging conditions. Brochothrix thermosphacta and lactic acid bacteria were found to be members of the final microbial flora of MAP and air-packaged mullet, whereas the Enterobacteriaceae population was lower than other bacterial groups. Of the chemical freshness indices determined, thiobarbituric acid values were variable in mullet samples irrespective of packaging conditions indicative of no specific oxidative rancidity trend. Based on sensorial data and aerobic plate count, trimethylamine nitrogen and total volatile basic nitrogen limit values in the range of ca. 15 to 23 and 52 to 60 mg N/100 g of fish muscle were obtained, respectively, for mullet packaged under modified atmosphere and air. Sensory analyses (odor and taste attributes) showed that the limit of sensorial acceptability was reached after ca. 6 days for the samples packaged in air, 8 days for the M1 and M3 samples, and after 10 days for the M2 samples. Respective shelf-life extension for fresh whole mullet was ca. 2 days (M1 and M3 gas mixtures), and 4 days (M2 gas mixture).

Animals↗
PubMed1993

RNA-mediated specificity of DNA packaging into hybrid lambda/phi 29 proheads.

A small RNA (pRNA, 174 nt) is known to be essential for DNA packaging in bacteriophage phi 29. However, in an in vitro DNA packaging system based on hybrid lambda/phi 29 proheads (made up of head proteins from phage lambda and connectors from phage phi 29), the specificity of DNA packaging is lost, and different RNA molecules fulfil the requirements for DNA packaging, albeit with less efficiency than phi 29 pRNA. Competition assays with RNAs from different sources have shown that phi 29 connectors bind preferentially pRNA. An increase in the efficiency of phi 29 DNA packaging into hybrid proheads induced by phi 29 pRNA is observed because, when phi 29 pRNA is incubated with hybrid proheads, phi 29 DNA is packaged more efficiently than other DNAs of similar length. Furthermore, when hybrid proheads carrying phi 29 pRNA are incubated with a mixture of DNAs from different sources, phi 29 DNA is selectively packaged, thus indicating that phi 29 pRNA determines the specificity of DNA packaging.

Amino Acid Sequence↗
PubMed2004

Production of lentiviral vectors by transient expression of minimal packaging genes from recombinant adenoviruses.

BACKGROUND: The potential of lentiviral vectors for clinical gene therapy has not yet been evaluated. One of the reasons is the cytotoxicity of lentiviral packaging genes which makes the generation of stable producer cell lines difficult. Therefore, a novel packaging system for lentiviral vectors based on transient expression of packaging genes by recombinant adenoviruses was developed. METHODS: Adenoviral vectors expressing VSV-G, codon-optimized HIV-1 gag-pol, and codon-optimized SIV gag-pol under the control of a tetracycline-regulatable promoter (adenoviral lenti-pack vectors) were constructed and the production levels of this vector system were evaluated. RESULTS: The generated adenoviral lenti-pack vectors could be grown to high titers when transgene expression was suppressed and no evidence for instabilities was obtained. Cells stably transfected with a SIV-based vector construct were converted into lentiviral vector producer cells by infection with the adenoviral lenti-pack vectors. Lentiviral vector titers obtained were as high as vector titers obtained by transient cotransfection experiments. A protocol was developed that allowed preparation of lentiviral vector stocks with undetectable levels of contaminating adenoviral lenti-pack vectors. CONCLUSIONS: The adenoviral lenti-pack vectors described should provide a convenient alternative approach to inducible packaging cell lines for large-scale lentiviral vector production. Transient expression of cytotoxic lentiviral packaging genes by the adenoviral lenti-pack vectors circumvents loss of titers during prolonged culture of packaging cell lines. The design of the adenoviral lenti-pack vectors should reduce the risk of transfer of packaging genes to target cells and at the same time provide flexibility with respect to the lentiviral vector constructs that can be packaged.

Adenoviridae↗
PubMed2000

In vitro packaging of DNA of the Bacillus subtilis bacteriophage SPP1.

In vitro packaging of bacteriophage SPP1 DNA into procapsids is described and the requirements of this process were determined. Combination of proheads with an extract supplying terminase, DNA and phage tails yielded up to 10(7 )viable phages per milliliter of in vitro reaction under optimized conditions. The presence of neutral polymers and polyamines had a concentration and type dependent effect in the packaging reaction. The terminase donor extract lost rapidly activity at 30 degrees C in contrast to the stability of the prohead donor extract. Maturation to infective virions was observed using both procapsids assembled in SPP1 infected cells and procapsid-like structures assembled in Escherichia coli that overexpressed the SPP1 prohead gene clusters. Neither a majority of aberrant capsid-related structures present in the latter material nor procapsids lacking the portal protein inhibited DNA packaging. Addition of purified portal protein reduced DNA packaging activity in vitro only at concentrations 20-fold higher than those found in the SPP1 infected cell. The SPP1 DNA packaged in vitro originated exclusively from the terminase donor extract. This packaging selectivity was not observed in vivo during mixed infections. The data are compatible with a model for processive headful DNA packaging in which terminase and DNA co-produced in the same cell are tightly associated and can effectively discriminate the portal vertex of DNA packaging-proficient proheads from aberrant structures, from portal-less procapsids, and from isolated portal protein.

Bacillus Phages↗
PubMed2001

Characterization of cis-acting sequences involved in canine adenovirus packaging.

The cis-acting packaging domain in adenovirus serotype 5 (Ad5) is a series of redundant, albeit not functionally equivalent, "A-repeats" made up of the consensus sequence 5'-TTTGN(8)CG-3'. A-repeats may bind trans-acting factors that direct packaging of the adenovirus genome into the preformed capsid. To try to understand this basic mechanism, we examined the packaging domain from a nonhuman adenovirus. We delimited the canine adenovirus type 2 (CAV-2) packaging domain to within 156 bp via a conditional mutation based on the Cre/loxP excision. Using an insertion, deletion, and substitution strategy, we generated packaging-defective CAV-2 vectors. Our results demonstrate that, like Ad5, CAV-2 cis-acting packaging sequences are located near the left inverted terminal repeat and are redundant, but not functionally equivalent. However, the bipartite motif found in Ad5 is present only once in CAV-2 and deletion of it caused only a minor variation in the packaging efficiency. We have identified at least four functional cis-acting packaging sequences in CAV-2. The CAV-2 vectors that we generated were not replication-defective in an E1-transcomplementing cell line and as heat stable as the parental vectors that did not contain mutations.

Adenoviridae↗
PubMed1995

A defined system for in vitro lambda DNA packaging.

We constructed a defined in vitro system for packaging lambda DNA which is composed of purified proheads, the noncapsid proteins terminase and gpFI, and the Escherichia coli DNA binding/bending protein IHF. The defined packaging system: (i) is free from endogenous ATP, DNAs, and DNases and (ii) packages 30% of the input mature lambda DNA efficiently. In this defined packaging system, IHF and gpFI make modest contributions to packaging efficiency. The defined packaging reactions showed that DNA packaging gave a linear response to the concentration of mature lambda DNA and terminase. DNA packaging showed a sigmoidal relationship with respect to the concentration of ATP and proheads.

Adenosine Triphosphate↗
PubMed1996

A novel human amphotropic packaging cell line: high titer, complement resistance, and improved safety.

Successful retroviral-mediated gene therapy will depend on safe, efficient packaging cell lines for vector particle production. Existing packaging lines for murine leukemia virus (MLV)-based vectors are predominantly derived from NIH/3T3 cells which carry endogenous MLV sequences that could participate in recombination to form replication-competent retrovirus (RCR). To identify cells devoid of such sequences, we screened genomic DNA from eight cell lines. DNA from the human 293 cell line did not cross-hybridize with MLV sequences, and these cells were able to secrete Gag particles after transfection. We derived a stable amphotropic packaging cell line (called ProPak-A) in 293 cells in which the Gag-Pol and Env (packaging) functions are expressed separately from a heterologous (non-MLV) promoter, to maximally reduce homology between packaging and vector sequences. ProPak-A-based producer cells are efficient, yielding higher stable titers than PA317-based producers. In addition, a vector that consistently gave rise to RCR in PA317 cells never resulted in detectable RCR in ProPak-A-based producer cultures. We have also shown that ProPak-A-packaged particles are not inactivated by human serum. Thus, the packaging cells we describe are as efficient and safer than the amphotropic packaging cells most commonly used in clinical gene therapy work and are also more appropriate for in vivo gene delivery.

3T3 Cells↗
PubMed2004

Helical packaging of semiflexible polymers in bacteriophages.

We investigate multilayered helical packaging of double-stranded DNA, or of a general polymer chain with persistence length lb, into an ideal, inert cylindrical container, reaching densities slightly below close packaging. We calculate the free energy as a function of the packaged length, based on the energies for bending, twisting, the suffered entropy loss, and the electrostatic energy in a Debye-Hückel model. In the absence of charges on the packaged polymer, a critical packaging force can be determined, similar to the mechanism involved in DNA unzipping models. When charges are taken into consideration, in the final packaging state the charges which are chemically distant become geometrically close, and therefore a steep rise is seen in the free energy. We argue that due to the extremely ordered and almost closely packaged final state the actual packaging geometry does not influence the behaviour of the free energy, pointing towards a certain universality of this state of the polymer. Our findings are compared to a recent simulations study, showing that the model is sensitive to the screening length.

Bacteriophage lambda↗
PubMed2000

Coordinate packaging of newly synthesized phosphatidylcholine and phosphatidylglycerol in lamellar bodies in alveolar type II cells.

Methylamine, a weak base, inhibits packaging of newly synthesized phosphatidylcholine (PC) in lamellar bodies in 20-22 h cultured alveolar type II cells, suggesting a role for acidic pH of lamellar bodies. In this study, we tested if (i) the packaging of PC is similarly regulated in freshly isolated type II cells and (ii) methylamine also inhibits the packaging of other surfactant phospholipids, particularly, phosphatidylglycerol (PG). The latter would suggest coordinated packaging so as to maintain the phospholipid composition of lung surfactant. During the short-term metabolic labeling experiments in freshly isolated type II cells, methylamine treatment decreased the incorporation of radioactive precursors into PC, disaturated PC (DSPC), and PG of lamellar bodies but not of the microsomes, when compared with controls. The calculated packaging (the percentage of microsomal lipid packaged in lamellar bodies) of each phospholipid was similarly decreased (approximately 50%) in methylamine-treated cells, suggesting coordinated packaging of surfactant phospholipids in lamellar bodies. Equilibrium-labeling studies with freshly isolated type II cells (as is routinely done for studies on surfactant secretion) +/- methylamine showed that in methylamine-treated cells, the secretion of PC and PG was decreased (possibly due to decreased packaging), but the phospholipid composition of released surfactant (measured by radioactivity distribution) was unchanged; and the PC content (measured by mass or radioactivity) of lamellar bodies was lower, but the PC composition (as percentage of total phospholipids) was unchanged when compared with control cells. We speculate that the newly synthesized surfactant phospholipids, PC, DSPC, and PG, are coordinately transported into lamellar bodies by a mechanism requiring the acidic pH, presumably, of lamellar bodies.

Ammonium Chloride↗
PubMed1988

Cloning, overexpression and purification of the terminase proteins gp16 and gp17 of bacteriophage T4. Construction of a defined in-vitro DNA packaging system using purified terminase proteins.

Terminases of double-stranded DNA bacteriophages are required for packaging and generation of terminii in replicated concatemeric DNA molecules. Genetic evidence suggests that these functions in phage T4 are carried out by the products of genes 16 and 17. We cloned these T4 genes into a heat-inducible cI repressor-lambda PL promoter vector system, and overexpressed them in Escherichia coli. We developed an in-vitro DNA packaging system, which, consistent with the genetic data, shows an absolute requirement for the terminase proteins. The overexpressed terminase proteins gp16 and gp17 appear to form a specific complex and an ATP binding site is present in the gp17 molecule. We purified the terminase proteins either as individual gp16 or gp17 proteins, or as a gp16-gp17 complex. The gp16 function of the terminase complex is dispensable for packaging mature DNA, whereas gp17 is essential for packaging DNA under any condition tested. We constructed a defined in-vitro DNA packaging system with the purified terminase proteins, purified proheads and a DNA-free phage completion gene products extract. All the components of this system can be stored at -90 degrees C without loss of packaging activity. The terminase proteins, therefore, may serve as useful reagents for mechanistic studies on DNA packaging, as well as to develop T4 as a packaging-cloning vector.

Cloning, Molecular↗