Christiansen B, Johnsen M G, Stenby E, Vogensen F K, Hammer K
Department of Microbiology, Technical University of Denmark, Lyngby.
J Bacteriol. 1994 Feb;176(4):1069-76. doi: 10.1128/jb.176.4.1069-1076.1994.
The temperate lactococcal phage TP901-1, induced by UV light from Lactococcus lactis subsp. cremoris 901-1, was characterized. The restriction map was found to be circular, and the packaging of TP901-1 DNA was concluded to occur by a headful mechanism. The pac region was localized on the 38.4-kb phage genome. TP901-1 belongs to the class of P335 phages (V. Braun, S. Hertwig, H. Neve, A. Geis, and M. Teuber, J. Gen. Microbiol. 135:2551-2560, 1989). Evidence is presented that the phages TP936-1 (V. Braun, S. Hertwig, H. Neve, A. Geis, and M. Teuber, J. Gen. Microbiol. 135:2551-2560, 1989) and C3-T1 (A. W. Jarvis, V. R. Parker, and M. B. Bianchin, Can. J. Microbiol. 38:398-404, 1992) are very closely related to or are identical to TP901-1. The lytically propagated TP901-1 phages were able to lysogenize both indicator strains Lactococcus cremoris 3107 and Wg2. Lysogenization resulted in site-specific integration of the phage genome into the bacterial chromosome. Only one chromosomal attB site was found in 20 independent lysogens. The attP region of TP901-1 and the attL and attR regions were cloned and sequenced. The results showed a core region of only 5 bp, in which the recombination occurs, followed after a 1-bp mismatch by a 7-bp identical region, TCAAT(T/C)AAGGTAA. This result was further verified by sequencing of the attB region obtained by PCR. An integration vector was constructed with the 6.5-kb EcoRI fragment from TP901-1 containing attP. This vector also functions in the plasmid-free strains, MG1363 and LM0230 with only one specific attB site, strongly indicating a more general use of the TP901-1-based integration vector in lactococci.
对由乳酸乳球菌亚种cremoris 901 - 1经紫外线诱导产生的温和型乳球菌噬菌体TP901 - 1进行了特性分析。发现其限制酶切图谱呈环状，并且得出TP901 - 1 DNA的包装是通过“满头部”机制进行的结论。pac区域定位在38.4 kb的噬菌体基因组上。TP901 - 1属于P335噬菌体类别（V. Braun、S. Hertwig、H. Neve、A. Geis和M. Teuber，《普通微生物学杂志》135:2551 - 2560，1989年）。有证据表明噬菌体TP936 - 1（V. Braun、S. Hertwig、H. Neve、A. Geis和M. Teuber，《普通微生物学杂志》135:2551 - 2560，1989年）和C3 - T1（A. W. Jarvis、V. R. Parker和M. B. Bianchin，《加拿大微生物学杂志》38:398 - 404，1992年）与TP901 - 1密切相关或相同。经裂解繁殖的TP901 - 1噬菌体能够使指示菌株乳酸乳球菌cremoris 3107和Wg2发生溶源化。溶源化导致噬菌体基因组位点特异性整合到细菌染色体中。在20个独立的溶源菌中仅发现一个染色体附着位点attB。对TP901 - 1的attP区域以及attL和attR区域进行了克隆和测序。结果显示仅有一个5 bp的核心区域发生重组，在一个1 bp错配之后是一个7 bp的相同区域，即TCAAT(T/C)AAGGTAA。通过对PCR获得的attB区域进行测序进一步验证了该结果。用来自TP901 - 1的包含attP的6.5 kb EcoRI片段构建了一个整合载体。该载体在无质粒菌株MG1363和LM0230中也能发挥作用，这两种菌株仅有一个特定的attB位点，这有力地表明基于TP901 - 1的整合载体在乳球菌中有更广泛的用途。
