Portable Color Doppler Ultrasound Series Color Doppler Ultrasound System
-Abundant function Multilingual
Specifications Portable Color Doppler Ultrasound,Portable Color Ultrasound Machine,Portable Color Ultrasound Equipment,Color Ultrasound Equipment For Clinic Guangzhou Sonostar Technologies Co., Limited , https://www.sonoeye.com
Second, processing genomic DNA
According to the research purpose, researchers need to choose the appropriate carrier. Different vectors have different requirements for the length of genomic DNA, and genomic DNA of a suitable length must be selected to construct a genomic library. For example, for a cosmid vector, a suitable fragment length is about 40 kb; for a BAC vector, a suitable fragment length is on average 120 kb to 300 kb.
To construct a cosmid genomic DNA library, the genomic DNA needs to be randomly slashed with a syringe; then repairing the DNA with a terminal repair enzyme can increase the efficiency of DNA ligation into the vector; then, by pulse field electrophoresis or ordinary electrophoresis, 40 kb can be found. The DNA fragment is then recovered using a gel recovery kit.
To construct a BAC genomic DNA library, the genomic DNA needs to be digested with restriction endonucleases (usually EcoR I, BamH I or Hind III), and then a suitable length of DNA fragments (such as 100 kb-150 kb) can be found by pulse field electrophoresis. The DNA is then recovered by dialysis.
requires attention:
(1) When electrophoresis, the appropriate DNA Ladder should be selected to ensure accurate positioning of the DNA of the desired molecular weight after electrophoresis.
(2) After electrophoresis, the time of irradiating the target DNA with ultraviolet light should be shortened as much as possible, otherwise the cloning efficiency will be significantly reduced.
(3) When recovering DNA, excessive centrifugation should be avoided to prevent DNA from being sheared and to reduce library quality.
Third, the choice of carrier
At present, commonly used genomic library vectors include a cosmid vector, a P1 phage vector, a PAC vector (P1 artificial chromosome), a BAC vector (bacterial artificial chromosome), and a YAC vector (yeast artificial chromosome). Which carrier to choose can refer to the following factors:
1, the size of the target area:
If the target region of the genome is small (less than 50 kb), a cosmid vector or even a lambda phage vector can be selected. A cosmid vector library can be conveniently constructed using existing commercial cosmid vectors, highly efficient packaging mixtures, and suitable E. coli strains.
If the target area of ​​the genome is large, then P1, PAC or BAC is more appropriate. P1 operation is difficult, and PAC is relatively simple. BAC vectors are also a good choice to build BAC libraries using existing mature products, such as a range of commercial BAC vectors and kits.
If the target area is very large (greater than 250kb), the YAC carrier is preferred. However, the use of YAC vectors to construct genomic libraries is very cumbersome to operate.
Table 1 Comparison of various carriers
2. The difficulty of screening the library: Carrier Capacity (kb) Host Introducing cells Clay 30-45 E.coli divert P1 70-100 E.coli divert PAC 130-150 E.coli Electric turn BAC 120-300 E.coli Electric turn YAC 250-400 yeast Conversion
Cosmid vectors are typically screened using conventional filter-printing hybridization, but this method is laborious and wasteful for constructing regional maps and contigs. Large-capacity vector libraries, such as P1, PAC, BAC, and YAC, are stored in a two-dimensional array, either by conventional single-copy probe hybridization or by PCR for polyclonal population screening. Moreover, using a high volume vector to construct a library can reduce the steps of chromosome walking.
3, the consideration of the number of copies of the carrier:
When a large fragment DNA fragment is cloned into a high-copy vector, the cloned DNA is recombined, thereby affecting the fidelity and stability of the cloned sequence; while the low-yield DNA of the single-copy vector is a bottleneck for high-throughput analysis. The solution to this conflict can be EPICENTRE's CopyControl TM cloning system, CopyControl TM technology combines the advantage of a single copy and multiple copy vector vector. The single-copy vector can improve the stability of the insert, and can simultaneously amplify a high copy number clone to obtain high-yield DNA under the action of an inducer.
4. Link the genomic DNA fragment into the vector
The above-described DNA fragment of the appropriate size is ligated into the vector using a ligase. Many commercial vectors have been pretreated and can be used directly without the need for endonuclease digestion and dephosphorylation. It is preferred to use a ligation enzyme with fast connection and high efficiency to connect the reaction system to 100 μl.
Note: After the BAC vector is connected, the ligation product needs to be desalted to remove the salt from the ligation reaction buffer.
5. Transfer of the recombinant vector into the host cell <br> If the cosmid library is constructed, the above-mentioned ligation reaction product is packaged with the packaging protein, and the titer of the packaged cosmid clone is determined, and then transfected. The recombinant is picked and the size of the insert is identified. If the size and quality of the library are satisfactory, the library can be screened, or the library can be expanded and saved.
If a BAC library is constructed, the electroporation method should be selected to transfer the above-mentioned ligation product to the recipient bacteria, and the plate is grown to clone. After obtaining the clone, the size of the BAC clone needs to be evaluated to determine whether the size of the library can meet the requirements. If the library is the right size, you can do the rest.
Determination of the number of library clones
The genomic DNA library needs to contain enough clones to ensure the representativeness of the library. Generally use the following empirical formula to determine:
N = ln (1-P ) / ln (1-f ) P is the desired coverage, f is the ratio of the insert size to the size of the genomic DNA, and N is the number of clones required.
For example, using the BAC vector to construct a human genomic library with a human genome size of 3 x 10 9 bp, an insert size of 100 kb, and a desired coverage of 99%, then the number of BAC clones required is
N = ln (1 - 0.99) / ln (1 - [10 6 bp / 3 x 10 9 bp]) = 138,298
-High definition image
-Very thin light and pretty
-15inch LED Screen
-Built-in Battery
1. Imaging Modes:
-B, B|B, 4B, B|M, M
-Color Doppler (CFM)
-Power Doppler (PDI)
-Directional Power Doppler (DPDI)
-Pulsed Wave Doppler (PWD)
-B+PWD (Duplex)
-B+CFM/PDI/DPDI+PWD (Triplex)
-High Pulse Repetition Frequency (HPRF)
-Tissue Harmonic Imaging (THI)
2. Scanning Method: electronic linear, electronic convex, electronic micro-convex, scanning depth: 2-24cm
3. Color Doppler:
-PRF variable: 0.5-9 kHz
-Wall filter settings: 3 steps (5%, %10%, 15% PRF)
-Angle steering for linear transducers: ±10°
-Real-time spatial filter: 4 values
-CFM palette>10 maps
-PDI palette>10 maps
-B/Color priority control
-Color threshold control
-CFM baseline control
-Doppler frequency selection
-Color frame averaging
-Transparent Color Mapping (TCM)
4. Pulsed Wave Doppler:
-PRF variable: 1-10 kHz
-Wall filter settings: 16 steps (2.5%-20% PRF)
-Angle steering for linear transducers: ±10°
-Real-time trace line with automatic calculation of spectrum parameters
-Stereo sound: volume control
-PWD palette>10 maps
-Doppler frequency selection
5.Processing:
-High Line Density scan mode for better resolution
-8 sliders TGC Control
-Dynamic range>120 dB
-Overall gain control
-M - mode sweep speed control
-Acoustic power control
-Variable frame averaging
-Brightness, contrast
-Advanced gamma control
-Scan direction, rotation, up-down controls
-Negative / positive control
-Echo enhancement control
-Noise rejection function
-Speckle reduction
6. Image and video: AVI, JPG, BMP, PNG, TIF, DCM (DICOM)
7. General Measurements and Calculations: Distance, Length, Area, Circumference, Volume, Angle, Stenosis %, A/B Ratio, Velocity, Pressure Gradient (PG), Acceleration, Resistivity Index (RI), Heart Rate, Velocity Time Integral (VTI), etc.
8. Measurements and Calculations Software Packages: Obstetrics, Gynecology, Abdominal, Urology, Endocrinology, Vascular, Cardiology, etc.
9. Expansion interfaces:
-VGA, TV Interface
-USB2.0 Interface
-RJ-45 Network interface
-Support DeskJet printer, LaserJet printer, video printer
Essential experimental techniques for genomic DNA library construction
Genomic DNA libraries are widely used, such as in human and animal and plant genomics research, gene expression regulation research, analysis, and isolation of specific gene fragments. In general, the basic flow of genomic library construction can be classified into four major steps: isolating genomic DNA, performing related processing on genomic DNA, ligating genomic DNA fragments into vectors, and transferring recombinant vectors into host cells.
Isolating Genomic DNA (gDNA) There is no doubt that high quality genomic DNA is critical for genomic library construction. It is necessary to select the appropriate genomic DNA isolation method through experience, to ensure that the DNA is not excessively sheared or degraded during the separation process, and to ensure the purity of the DNA as much as possible.