Agrobacterium tumefaciens can transfer a DNA fragment (T-DNA) from its Ti plasmid to host plant and integrate the T-DNA fragment into host cell nuclear genome. Agrobacterium-mediated T-DNA transfer is the most widely used genetic transformation method for plant. The T-DNA is delivered in the form of single-stranded T-DNA-protein complex (T-complex) by the polar-located Agrobacterium type Ⅳ secretion system (T4SS) to the host cell. T4SS is ancestrally related to bacterial conjugation machines and still used by many plasmids as conjugation channel. Moreover, T4SS is also the secretion channel that used by many human bacterial pathogens to inject the effector proteins to host cells, thus contributing directly to the bacterial pathogenicity. Therefore, in addition to the technical application as a gene vector to create transgenic plants, Agrobacterium-mediated T-DNA transfer also provides a fascinating model system to study the intercellular transfer of macromolecules. The study on the molecular mechanism of T-DNA transfer arouses extensive interest and progresses rapidly in recent years. Here the recent advances in research on T-complex formation and T-complex transfer in Agrobacterium cell are reviewed.

Agrobacterium tumefaciens possesses many advantages as a model bacterium for the study of a wide variety of biological processes. Gene disruption or inactivation is a powerful and direct tool for investigation of in vivo gene functions. The intensive study ofA. tumefaciens has increased the need for simple and highly efficient procedures to manipulate its genome. The sacB gene was used as a counterselectable marker to develop a gene replacement procedure that allows precise insertion, deletion, and allele substitution of any gene sequence in A. tumefaciens without altering the genome in any other way. A kanamycin resistance (KmR) cassette was constructed to the suicide vector as the positive selection marker. The suicide plasmid containing DNA fragments homologous to the flanking sequences of the target gene was integrated into the recipient cell genome at the target gene locus by intermolecular homologous recombination, generating the KmR-single cross-over colonies. The effect of homologous sequence length on the intermolecular homologous recombination was analyzed. The second cross-over colonies generated by intramolecular homologous recombination occurring between two tandem repeats were simply screened out by counter-selection of sacB. Data showed that the intervening sequence length between two repeats significantly affected the intramolecular homologous recombination frequency in A. tumefaciens, indicating that A. tumefaciens adopted the homologous recombination mechanism similar to that in E. coli. All these results demonstrated that investigators could minimize the numbers of colonies to be analyzed and reduce the overall workload by optimizing the relative length of two homologous fragments and using the specific type of single cross-over transformants for screening the second cross-over event. This mutagenesis strategy had successfully been used to generate the double unmarked △vbp2△vbp3 mutant in two A. tumefaciens strains.

Objective:To describe the COVID-19 epidemic and its characteristics in Heilongjiang province, and provide evidence for the further prevention and control of COVID-19 in the province.Methods:The information of COVID-19 cases and clusters were collected from national notifiable disease report system and management information system for reporting public health emergencies of China CDC. The Software’s of Excel 2010 and SPSS 23.0 were applied for data cleaning and statistical analysis on the population, time and area distributions of COVID-19 cases.Results:On January 22, 2020, the first confirmed case of COVID-19 was reported in Heilongjiang. By March 11, 2020, a total of 482 cases domestic case of COVID-19, The incidence rate was 1.28/100 000, the mortality rate was 2.70% (13/482) in 13 municipalities in Heilongjiang. There were 81 clusters of COVID-19, The number of confirmed cases accounted for 79.25% (382/482) of the total confirmed cases and 12 cases of deaths. The family clusters accounted for 86.42% (70/81). Compared with the sporadic cases, the mortality rate, proportion of elderly cases aged 60 or above and severe or critical cases of clinical classification were all higher in the clusters especially the family clusters, but the differences were not significant ( P>0.05). There were 34 clusters involving more than 5 confirmed cases accounted for 41.98% (34/81) of the total clusters, the involved cases accounted for 68.31% (261/382) of the total cases of clusters. There were significant differences in age distribution of the cases among the case clusters with different case numbers. In the clusters involving 6-9 cases, the proportion of cases aged 65 years or above was more (26.53%, 39/147). Conclusions:The incidence rate of COVID-19 was relatively high and the early epidemic was serious in Heilongjiang, The number of cases was large in clusters especially family clusters.