Identification of positive GATEWAY expression clones when both the pENTRY and pDEST vectors contain the same marker for bacterial selection (Record no. 54392)

MARC details
000 -LEADER
fixed length control field 02661nam a2200181Ia 4500
003 - CONTROL NUMBER IDENTIFIER
control field MX-MdCICY
005 - DATE AND TIME OF LATEST TRANSACTION
control field 20250625162446.0
040 ## - CATALOGING SOURCE
Transcribing agency CICY
090 ## - LOCALLY ASSIGNED LC-TYPE CALL NUMBER (OCLC); LOCAL CALL NUMBER (RLIN)
Classification number (OCLC) (R) ; Classification number, CALL (RLIN) (NR) B-20293
008 - FIXED-LENGTH DATA ELEMENTS--GENERAL INFORMATION
fixed length control field 250602s9999 xx |||||s2 |||| ||und|d
245 10 - TITLE STATEMENT
Title Identification of positive GATEWAY expression clones when both the pENTRY and pDEST vectors contain the same marker for bacterial selection
490 0# - SERIES STATEMENT
Volume/sequential designation Cold Spring Harbor Protocols, 6, p.pdb. prot4647.4647, 2006
520 3# - SUMMARY, ETC.
Summary, etc. GATEWAY cloning technology (Invitrogen)takes advantage of bacteriophage ? site-specific recombination. The life cycle of ? alternates between the lytic and lysogenic stages. DNA can be inserted or excised from the Escherichia coli host genome by recombination between specific sites, AttB (bacterial)and AttP (phage). This process is mediated by the ? proteins int (integrase)and xis (excisionase), and a host protein IHF (integration host factor). GATEWAY cloning technology uses this process to insert fragments of DNA directionally into specially adapted vectors. These vectors contain a negative selectable marker, the ccdB gene, to select against nonrecombinant clones. Promoter or gene fragments are made GATEWAY compatible with adapter primers and amplified by PCR. These fragments are used in a BP clonase reaction to create ENTRY clones. Usually the pDONR vector used to generate such ENTRY clones is chosen so that the antibiotic selection marker is different from that of the pDEST vector, which finally generates an expression clone. This favors the selection of the expression clone and selects against the pENTRY clone. Now that many pENTRY and pDEST vectors have been generated and made available in stock centers, the antibiotic resistance genes are predetermined and may not always be compatible with each other. This problem is frequently experienced by plant researchers, since many full-length cDNA libraries have been generated using the pDONR-TOPO, pDONR221, or pENTR1A vectors, which are all kanamycin resistant in bacteria, and many pDEST vectors have been adapted from conventional plant transformation vectors, which are also frequently kanamycin resistant in bacteria. The following protocol describes ways in which such difficult vector combinations can be used effectively to obtain the appropriate expression clone without having to convert the pENTRY clone or pDEST clone to vectors with compatible antibiotic resistances.
700 12 - ADDED ENTRY--PERSONAL NAME
Personal name Basherudin, N.
700 12 - ADDED ENTRY--PERSONAL NAME
Personal name Curtis, M. D.
856 40 - ELECTRONIC LOCATION AND ACCESS
Uniform Resource Identifier <a href="https://drive.google.com/file/d/1GpXik5-SAG4IvMA4hy4nHYPhrJCaowmI/view?usp=drivesdk">https://drive.google.com/file/d/1GpXik5-SAG4IvMA4hy4nHYPhrJCaowmI/view?usp=drivesdk</a>
Public note Para ver el documento ingresa a Google con tu cuenta: @cicy.edu.mx
942 ## - ADDED ENTRY ELEMENTS (KOHA)
Source of classification or shelving scheme Clasificación local
Koha item type Documentos solicitados
Holdings
Lost status Source of classification or shelving scheme Damaged status Not for loan Collection Home library Current library Shelving location Date acquired Total checkouts Full call number Date last seen Price effective from Koha item type
  Clasificación local     Ref1 CICY CICY Documento préstamo interbibliotecario 25.06.2025   B-20293 25.06.2025 25.06.2025 Documentos solicitados