Abstract:
This invention relates in general to characterizing hydrocarbon reservoirs and/or determining flow properties of fluids associated with the reservoir-including fluids introduced into the reservoir to provide for hydrocarbon extraction-using biological tags and real-time polymerase chain reactions for tag detection. In embodiments of the present invention, one or more biological tags may be added to one or more liquids associated with the hydrocarbon reservoir and subsequently one or more liquid samples may be taken from locations associated with the hydrocarbon and the presence of the one or more biological tag may be qualitatively and/or quantitatively tested for in the samples using real-time PCR.

Description:
BACKGROUND OF THE INVENTION 
       [0001]    Embodiments of the present invention relate to characterization of hydrocarbon reservoirs and/or determining flow properties of fluids associated with such reservoirs. More specifically, but not by way of limitation, embodiments of the present invention provide for using biological tags as tracers that may be detected and/or measured in fluids retrieved from the hydrocarbon reservoir, one or more wellbores associated with the hydrocarbon reservoir or one or more locations proximal to the hydrocarbon reservoir using a real-time polymerase chain reaction (“real-time PCR”). From the detection and/or measurement of the biological tags in the retrieved fluids, the reservoir may be characterized and/or flow properties of fluids associated with the hydrocarbon reservoir may be determined. The fluids associated with the hydrocarbon reservoir may be fluids existing in the hydrocarbon reservoir and/or adjacent earth formations or may be fluids introduced into the hydrocarbon reservoir. 
         [0002]    In the specification, the term “real-time PCR” may refer to the monitoring of the amplification process in a polymerase chain reaction as the polymerase chain reaction proceeds. In certain aspects, the monitoring of the polymerase chain reaction may be provided by monitoring amplicon produced in the polymerase chain reaction, and such monitoring may be performed during one or more of the cycles of the polymerase chain reaction. 
         [0003]    Previously, characterization of reservoirs in earth formations or flow properties of fluids through earth formations has been provided for by the use of radioactive or chemical tracers. Traditional radioactive tracers include: the radionuclides 57Co, 58Co and 60Co (that may be incorporated into anionic complexes, such as hexacyanocobaltates etc.); 134Cs, 137Cs (as a corresponding chloride); tritiated water; and 22Na as sodium chloride. The most common beta emitting radioactive tracers for inter-well studies may be labelled with tritium (tritiated water) and carbon-14 (labelled thiocyanate). In radioactive tracer techniques, liquid scintillation counting may be used to detect the tracers found in fluid samples taken from earth formations or proximal locations. 
         [0004]    With regard to chemical tracers, some of the chemicals that have been used for reservoir characterization include chloride, bromide, iodide, nitrate, thiocyanate, fluorescein, rhodamine, 2-propanol, t-butanol and other water-soluble small organic molecules. In such techniques, after sample collection, detection of the chemicals may be conducted by techniques such as ion chromatography (for multiple anions), high-performance liquid chromatography, ultraviolet spectroscopy (for thiocyanate), colorimetry/fluorimetry (for dyes), conductivity (for ionic species), gas-liquid chromatography (for small organics). With chemical tracing, the detection limit—along with an estimate of the reservoir pore volume and expected dilution—determines the overall quantity of the chemical to be introduced into the hydrocarbon reservoir. Since a better description of the reservoir properties is the object of a tracer study, there may be a need to use large amounts of chemicals, which may be expensive and raise ecological/waste management issues. As such, the amount of tracer used is a balance between ensuring a measurable signal at the monitoring point(s) but with reasonable cost. 
         [0005]    Both radioactive and chemical tracer techniques, in practice, have many limitations, including that radioactive tracers require specialized environmental measures/precautions and chemical tracers require application of large doses of chemicals to provide for accurate detection and quantitative analysis. Moreover, with both radioactive and chemical tracers separate detection tests needs to be performed when more than one type of radioactive or chemical tracer is applied to the hydrocarbon reservoir. Furthermore, with regard to oil and gas reservoirs, processes such as hydraulic fracturing, hydrocarbon recovery processes and the like may have several stages and each stage of the process may have a fluid associated with the stage, including in fracturing, for example, pre-pad fluids, pad fluids, fracturing fluids and tailing fluids. Although these procedures may now be established in the oil and gas industry, there still exists significant room for improvement to the processes that may require understanding movements of the different fluids through the hydrocarbon reservoir and adjacent formations. 
       BRIEF SUMMARY OF THE INVENTION 
       [0006]    Embodiments of the present invention provide methods and systems for characterizing a hydrocarbon reservoir and/or determining flow characteristics of one or more liquids injected into and/or flowing in the hydrocarbon reservoir using biological tags and a real-time polymerase chain reaction. In one embodiment of the present invention, characterization of a hydrocarbon reservoir or determining flow properties of fluids associated with the hydrocarbon reservoir may be provided by adding a biological tag to a liquid injected into and/or associated with the hydrocarbon reservoir, collecting a fluid sample from the hydrocarbon reservoir, performing real-time PCR on the fluid sample to detect a presence of and/or measure an amount of the biological tag in the fluid sample, and using results from the real-time PCR to provide for at least one of the characterization of the hydrocarbon reservoir and determining flow properties of the liquid injected into and/or associated with the hydrocarbon reservoir. 
         [0007]    More specifically, but not by way of limitation, in embodiments of the present invention: (a) biological tags may be added to one or more liquids associated with the hydrocarbon reservoir, such as liquids injected into, liquids found in or flowing through the reservoir; (b) samples may be collected from the hydrocarbon reservoir or surrounding locations and multiple samples may be taken at different geographical or temporal locations; (c) the presence of the biological tag may be qualitatively and/or quantitatively tested for in the samples using real-time PCR; and (d) from the qualitative and/or quantitative results of the real-time PCR analysis the hydrocarbon reservoir may be characterized, the flow of the one or more liquids in the hydrocarbon reservoir may be mapped and/or where the one or more liquids has been added to the hydrocarbon reservoir—for a purpose such as fracturing, hydrocarbon recovery or the like—an analysis of the likely results of the fracturing process, hydrocarbon recovery or the like may be determined. 
         [0008]    In one embodiment, a hydrocarbon reservoir is characterized and/or flow properties of fluids in the hydrocarbon reservoir may be determined by injecting a first liquid containing a first biological tag into the hydrocarbon reservoir or adding the first biological tag to a liquid associated with the hydrocarbon reservoir, collecting a first fluid sample from the hydrocarbon reservoir, performing real-time PCR on the first fluid sample using a first pair of complementary primers in a polymerase chain reaction (“PCR”), wherein the first pair of complementary primers are configured to anneal to single strands of the first biological tag and provide for replication of the first biological tag during the real-time PCR, and wherein the step of performing real-time PCR comprises detecting PCR product produced by the replication of the first biological tag in the PCR reaction and using results from the real-time PCR to provide for at least one of characterization of the hydrocarbon reservoir and determining flow of the first liquid in the hydrocarbon reservoir. 
         [0009]    In certain aspects of the above embodiment, the biological tag may be DNA sequences and the first pair of complementary primers may be DNA fragments having DNA sequences complementary to the ends of the DNA sequence of the biological tag. In an embodiment of the present invention, the real-time PCR process may involve the use of probes configured to provide a detectable physical property that varies in accordance with amount of the PCR product, amplicon, produced during the real-time PCR. In such an embodiment, a value of the physical property may be measured to determine the presence of the PCR product and/or a relative quantity of the PCR product present. The PCR product may be the biological tags originally present in the sample undergoing real-time PCR, copies of the original biological tags generated by the PCR process and incomplete copies of the biological tags being generated in the PCR—i.e., the denatured single strands of the biological tags combined with the primers and/or nucleotide bases that are in the process of forming molecules equivalent to the biological tags. 
         [0010]    In certain aspects of embodiments of the present invention, the biological tags may be introduced into the hydrocarbon reservoir via a wellbore. In such aspects, the biological tags may be introduced into the hydrocarbon reservoir in a fracturing fluid, injection water, drilling fluid, tracer fluid and/or the like. Coiled tubing drilling techniques may be used in some aspects to provide for delivery of the biological tags into the hydrocarbon reservoir, a wellbore in the hydrocarbon reservoir, an earth formation proximal to the hydrocarbon reservoir and/or the like. 
         [0011]    In another embodiment of the present invention, a method for characterizing a hydrocarbon reservoir and determining flow of a plurality of fluids in the hydrocarbon reservoir is provided that comprises:
       injecting a first liquid containing first biological tags into the hydrocarbon reservoir;   injecting a second liquid containing second biological tags into the hydrocarbon reservoir;   collecting a first fluid sample from the hydrocarbon reservoir;   performing real-time PCR on the first fluid sample using a real-time PCR mixture, wherein the real-time PCR comprises:
           using a first primer to amplify any of the first biological tags present in the first fluid sample, wherein the first primer is configured to selectively attach to the first biological tags and provide for amplification of the first biological tags;   using a second primer to amplify any of the second biological tags present in the first fluid sample, wherein the second primer is configured to selectively attach to the second biological tags and provide for amplification of the second biological tags;   using a first probe to selectively detect the presence of the first biological tags, wherein the first probe is configured to produce a first measurable physical property that varies in accordance with a first amount of the first biological tags present in the real-time PCR mixture;   using a second probe to selectively detect the presence of the second biological tags, wherein the second probe is configured to produce a second measurable physical property that is distinct from the first measurable physical property and that varies in accordance with a second amount of the second biological tags-present in the real-time PCR mixture; and   
           using results from the real-time PCR to determine flow properties of the first and the second fluids in the hydrocarbon reservoir.       
 
         [0021]    In such an embodiment, the first and second liquids may be injected into the hydrocarbon reservoir through a borehole penetrating the hydrocarbon reservoir. 
         [0022]    In a further embodiment of the present invention, a system for characterizing a hydrocarbon reservoir and determining flow of fluids in the hydrocarbon reservoir is provided that may comprise a first well-tool configured to pump a first liquid containing a biological tag into the hydrocarbon reservoir, a sampling chamber configured to collect a fluid sample from the hydrocarbon reservoir, a real-time PCR device configured to perform real-time PCR of the fluid sample and to detect/measure the biological tag, and a processor configured to process outputs from the real-time PCR device to determine at least one of characterization of the hydrocarbon reservoir and flow of the first liquid in the hydrocarbon reservoir. 
         [0023]    Reference to the remaining portions of the specification, including the drawings and claims, will realize other features and advantages of the present invention. Further features and advantages of the present invention, as well as the structure and operation of various embodiments of the present invention, are described in detail below with respect to the accompanying drawings. 
     
    
     
       BRIEF DESCRIPTION OF THE DRAWINGS 
         [0024]    The present invention will become more fully understood from the detailed description and the accompanying drawings, wherein: 
           [0025]      FIG. 1A  is a schematic-type illustration of a wellbore penetrating a hydrocarbon reservoir and an associated wellbore assembly for introducing fluids containing biological tags into a hydrocarbon reservoir, in accordance with an embodiment of the present invention; 
           [0026]      FIG. 1B  is a schematic-type illustration of a sampling well-tool suspended in a wellbore penetrating a hydrocarbon reservoir configured for sampling fluids associated with the hydrocarbon reservoir and an associated real-time polymerase chain reaction processing station, in accordance with an embodiment of the present invention: 
           [0027]      FIG. 2A  is a flow type schematic illustrating a conventional polymerase chain reaction process; 
           [0028]      FIG. 2B  is a flow type schematic illustrating a real-time polymerase chain reaction process for analyzing samples of fluids associated with a hydrocarbon reservoir, in accordance with an embodiment of the present invention; and 
           [0029]      FIG. 3  is a flow type schematic illustrating a method for characterizing a hydrocarbon reservoir and/or determining flow properties of one or more liquids associated with the hydrocarbon reservoir, in accordance with an embodiment of the present invention. 
       
    
    
       [0030]    In the appended figures, similar components and/or features may have the same reference label. Further, various components of the same type may be distinguished by following the reference label by a dash and a second label that distinguishes among the similar components. If only the first reference label is used in the specification, the description is applicable to any one of the similar components having the same first reference label irrespective of the second reference label. 
       DETAILED DESCRIPTION OF THE INVENTION 
       [0031]    Specific details are given in the following description to provide a thorough understanding of the embodiments. However, it will be understood by one of ordinary skill in the art that the embodiments may be practiced without these specific details. For example, circuits may be shown in block diagrams in order not to obscure the embodiments in unnecessary detail. In other instances, well-known circuits, processes, algorithms, structures, and techniques may be shown without unnecessary detail in order to avoid obscuring the embodiments. 
         [0032]    Also, it is noted that the embodiments may be described as a process which is depicted as a flowchart, a flow diagram, a data flow diagram, a structure diagram, or a block diagram. Although a flowchart may describe the operations as a sequential process, many of the operations can be performed in parallel or concurrently. In addition, the order of the operations may be re-arranged. A process is terminated when its operations are completed, but could have additional steps not included in the figure. A process may correspond to a method, a function, a procedure, a subroutine, a subprogram, etc. When a process corresponds to a function, its termination corresponds to a return of the function to the calling function or the main function. 
         [0033]      FIG. 1A  is a schematic-type illustration of a wellbore penetrating a hydrocarbon reservoir and an associated wellbore assembly for introducing fluids into a hydrocarbon reservoir, in accordance with an embodiment of the present invention. Referring now to  FIG. 1 , a truck winch  10  or the like may be used to wind or unwind a wireline  15  or the like into and out of a wellbore  20  that penetrates a hydrocarbon reservoir  25 . The wireline  15  may be coupled with a well-tool  30  and provide for the movement of the well-tool  30  in the wellbore  20 , which may include movement of the well-tool  30  to locations within the hydrocarbon reservoir  25 . Positioning of the well-tool  30  in the wellbore  20  may be provided by a positioning wheel  35  or the like, which may be configured to maneuver the well-tool  30  in the wellbore  20 . 
         [0034]    In some embodiments of the present invention a pump  40  or the like may be used to introduce fluids into the hydrocarbon reservoir  25 . Fluids may be pumped directly into the wellbore  20  or may be introduced into the hydrocarbon reservoir  25  by the well-tool  30 , which may be coupled with the pump  40 . As persons of skill in the art are aware, perforations may be made in the casing of the wellbore  20  to provide for the application of fluids to different formation locations adjacent to the wellbore  20 . Further, methods of sealing off parts and/or sections of the wellbore  20  and plugging the perforations after introduction of fluids may provide for more precise, forceful and/or economical delivery of fluids to formations adjacent to the wellbore  20 . Using the described methods of entering a fluid into the hydrocarbon reservoir, in an embodiment of the present invention, a fluid containing biological tags may be introduced into the hydrocarbon reservoir. 
         [0035]    In certain aspects, coiled tubing techniques may be used as a means to provide for application/retrieval of a fluid containing the biological tags into/from the hydrocarbon reservoir  25  and/or an adjacent location. Coiled tubing is a long, continuous string of tubing, generally made of steel that may be used in the drilling of a borehole. Coiled tubing is increasing in popularity as a method of conducting operations in an oil or gas wellbore. Historically, drilling pipe was used for drilling and conducting operations inside a wellbore, usually several hundred or thousand feet under the surface of the ground. However, drill pipe must be assembled in sections and lowered into the wellbore over a long time period of many hours or days. Coiled tubing has emerged as one solution to this matter by providing a relatively fast and reliable method of conducting operations downhole within a wellbore, without using heavy and cumbersome jointed drilling pipe. Coiled tubing is used as a continuous strand, and therefore is easier and faster to use in many wellbore operations. Technological developments, improved service reliability, and the need to drive down industry costs have contributed to expanded uses for coiled tubing. 
         [0036]    In certain embodiments, a coiled tube borehole may be provided adjacent to and/or within another borehole associated with the hydrocarbon reservoir  25  and/or into or adjacent to the hydrocarbon reservoir  25 . As such, a borehole created by a coiled tubing drilling technique may be used to provide for introduction/injection of the biological tags to desired locations associated with the hydrocarbon reservoir  25  and/or sampling of fluids at locations associated with the hydrocarbon reservoir  25 . 
         [0037]    The fluid containing the biological tags and being added to and/or injected into the hydrocarbon formation may be a treatment fluid—i.e., a fluid configured to provide for such things as stimulation, isolation or control of reservoir gas or water—a fluid for use in a hydraulic fracturing process—such as a pre-pad fluid, a pad fluid, a fracturing fluid, a tailing fluid or the like—and/or the like. Alternatively or in combination, in certain embodiments of the present invention, biological tags may be added to a fluid already in or flowing through the hydrocarbon reservoir to determine flow properties for the fluid, determine changes to the fluids flow during extraction of hydrocarbons from the reservoir and/or the like. Such fluids may include formation liquid hydrocarbons, water and/or the like. 
         [0038]      FIG. 1B  is a schematic-type illustration of a sampling well-tool suspended in a wellbore penetrating a hydrocarbon reservoir configured for sampling fluids associated with the hydrocarbon reservoir and an associated real-time PCR processing station, in accordance with an embodiment of the present invention. As illustrated, in an aspect of the present invention a sampling tool  31  may be entered into the hydrocarbon reservoir  25  via the wellbore  20 . In such an aspect, the sampling tool  31  may be used to collect fluid samples from inside the wellbore  20 . In other aspects, fluid samples may be collected from locations including one or more of the following or the like: locations adjacent to the hydrocarbon reservoir  25  that may be inside or outside an earth formation; different locations inside the wellbore  20 ; a sampling borehole penetrating the hydrocarbon reservoir  25 , a sampling borehole penetrating an earth formation outside of the reservoir  25 . In some embodiments of the present invention, fluid samples may be collected at different geographical and or temporal locations. In such embodiments, the plurality of samples may be analyzed using real-time PCR and the results may be processed along with geographical and temporal data to provide for fluid flow analysis and/or reservoir characterization. 
         [0039]    In an embodiment of the present invention, the collected samples of the fluid may be taken to a real-time PCR processing station  50 . The real-time PCR processing station  50  may be at the wellhead, an onsite laboratory, an offsite laboratory or the like. The real-time PCR processing station may contain a real-time PCR processor, related computer systems, fluid analyzers and/or the like. At the real-time PCR processing station  50 , real-time PCR analysis of the collected samples may be performed to determine fluid flow type properties of fluids introduced into the wellbore  20 , associated with the hydrocarbon reservoir  25 , associated with adjacent earth formations (such as formation water) and/or the like. The processing station  50  may be networked with one or more pieces of equipment associated with the wellhead, wellbore and/or the reservoir and may provide for adjusting controls of the one or more pieces of equipment in accordance with results obtained from the real-time PCR analysis of the one or more of the fluid samples. 
         [0040]      FIG. 2  illustrates a conventional PCR process. In step  100  of the conventional PCR process, reactants may be mixed together as a PCR reaction mixture  101  in a PCR test tube  105 , which may be a test tube, vial, reaction vessel and/or the like. The PCR reaction mixture  101  of the conventional process may comprise the DNA to be amplified (in embodiments of the the present invention this may be the biological tags or equivalents), a DNA polymerase enzyme, small primer sequences of DNA, a supply of nucleotide bases and/or magnesium chloride. 
         [0041]    In step  110  of the conventional PCR process, the PCR reaction mixture  101  may be heated. This heating may occur in a specialized PCR machine designed to provide for performing the conventional PCR process. In certain aspects, the reaction mixture  101  may be heated to between 90-100 degrees Celsius or the like and this heating may take place for periods in the region of thirty seconds. At temperatures in excess of about 94 degrees Celsius, complete DNA strands  111  present in the PCR reaction mixture  101  may separate into single DNA strands  112  as the hydrogen bonds holding them together break down. 
         [0042]    In step  120  of the conventional PCR process, the PCR reaction mixture  101  containing the separated DNA strands  112  mixture may be cooled down. Cooling of the temperature of the reaction mixture  101  in this step may be to temperatures in the range of about 50 to 60 degrees Celsius. At such a temperature range, primers  121  contained in the reaction may bind/anneal to the single DNA strands  112 . The primers  121  may comprise short sequences of nucleotide bases which join to the beginning of the single DNA strands  112  to provide for the copying/amplification process to start. The primers  121  may join to the beginning of the separated DNA strands to provide for the full copying process to start. Busing complementary pairs of primers, it may be provided that the areas on either side of the target sequence are extended. 
         [0043]    The primers  121  have DNA sequences complementary to areas adjacent to each side of the target sequence. As such, if the DNA sequence around the region selected to be amplified is known, the correct primers may be chosen to provide for amplification of only the selected DNA sequence or maybe very close variants. In embodiments of the present invention, because the DNA sequences of the biological tags applied to the hydrocarbon reservoir are known/chosen, primers may be selected that will only provide for amplification of DNA fragments with the same DNA sequence as the biological tags and the biological tags may be selected with DNA sequences that are unlikely to occur “naturally” in the geological formation, hydrocarbon reservoir or fluid being analyzed. As such, embodiments of the present invention may be very accurate in detecting small amounts of the biological tags being used as tracers with a very low risk of contamination from DNA fragments naturally occurring in the hydrocarbon reservoir. The primers  121  may be constructed in a laboratory or purchased from commercial suppliers. 
         [0044]    In step  130  of the conventional PCR process, the PCR reaction mixture  101  may be heated up to a temperature in the range of about 70 to 80 degrees Celsius. At this temperature, DNA polymerase enzyme  131  may act to add bases to the primers  121  so as to build up, as illustrated by arrows  133   a  and  133   b,  strands of DNA complementary to the single DNA strands  112  so as to form complete molecules/DNA fragments that are identical to the molecules/DNA fragments of the original DNA molecules/fragments that were provided in the PCR test tube  105  in step  100  to be amplified/copied. Steps  110 ,  120  and  130  may be repeated in what are known as cycles to provide for further amplification/copying of the selected DNA. By repeating steps  110 ,  120  and  130  around thirty times, it is possible to produce in the order of 1 billion copies of the original DNA selected for amplification/copying. However, such processing may take in the order of several hours, and, in conventional PCR is only a qualitative process, not quantitative, in that it only establishes the presence of a certain DNA in the PCR reaction mixture  101 , not how much of the certain DNA was originally present in the PCR test tube  105 . 
         [0045]      FIG. 2B  is a flow type schematic illustrating a real-time PCR process for analyzing fluids associated with a hydrocarbon recovery process, in accordance with an embodiment of the present invention. In an embodiment of the present invention, as discussed in more detail previously, biological tags may be introduced into a fluid associated with a hydrocarbon reservoir to act as tracers. Fluid samples may then be sampled from the reservoir or areas proximal to the reservoir to test for the biological tags and, as a result, to determine flow properties of the fluid and/or characterization of the hydrocarbon reservoir. 
         [0046]    In an embodiment of the present invention, in step  150 , a sample—the sample having been taken from the hydrocarbon reservoir, a wellbore associated with the reservoir or a location associated with the hydrocarbon reservoir after introduction of biological tags into one or more fluids associated with the hydrocarbon reservoir—is placed in a test tube  105  (which may be a vial, chemical container or the like) and mixed with a real-time PCR reaction mixture  151  comprising a DNA polymerase enzyme, primer sequences of DNA, a supply of nucleotide bases, some real-time PCR probes and/or the like. 
         [0047]    As discussed above, the biological tags and the primers may be selected so as to be complimentary to provide for selective copying/amplification of only any DNA fragments in the fluid sample that have the same DNA sequence as the biological tags. This use of primers specific to the biological tags introduced into the fluid associated with the hydrocarbon reservoir may provide for accurately detecting low levels of the DNA fragments because, among other reasons, there is little background noise from detections of similar DNA fragments and probabilities that DNA fragments matching the DNA fragments introduced into the fluid may occur in the sample from a source other than the deliberate introduction for flow analysis, as described above, are very low. 
         [0048]    The real-time PCR probes in the real-time PCR reaction mixture  151  may comprise binding dyes—such as SYBR® Green—hybridization or hydrolysis probes (that may be fluorescently labeled sequence-specific probes) such as QuantiProbes®, TaqMan® probes, FRET probes, molecular beacons and/or the like. Further, in certain aspects, the real-time PCR reaction mixture  151  may comprise primers/probes such as Scorpion™ probes, Sunrise™ primers, LUX™ fluourogenic primers and/or the like. 
         [0049]    In step  160 , the real-time PCR reaction mixture  151  may be heated. In certain aspects the real-time PCR reaction mixture  151  may be heated to a temperature above about 94 degree Celsius or the like to provide for denaturing of any DNA fragments  161  in the real-time PCR reaction mixture  151  into single DNA fragment strands  162  as the hydrogen bonds holding the DNA strands together break down. 
         [0050]    In step  170 , the real-time PCR reaction mixture  151  containing the single DNA fragment strands  172  may be cooled down. Cooling of the temperature of the real-time PCR reaction mixture  151  in this step may be to temperatures in the range of about 50 to 60 degrees Celsius. At such a temperature range, any primers in the reaction may bind/anneal to any single DNA fragment strands. In an embodiment of the present invention, primers  171  may comprise short sequences of nucleotide bases which may be specifically selected to join to an end of single tracer DNA fragment strands  172  (where the single tracer DNA fragment strands  172  are formed from the denaturing in step  160  of the biological tags) to provide for the copying/amplification of the single tracer DNA fragment strands  172 . In this way the primers  171  may provide for selective amplification/copying of only the single tracer DNA fragment strands  172 , and in turn the selective amplification of the biological tags in the sample. 
         [0051]    In certain embodiments of the present invention, more then one biological tag may be introduced into the hydrocarbon reservoir or fluids associated with the hydrocarbon reservoir. For example, multiple liquids may be used in a fracturing process performed on the hydrocarbon reservoir and/or surrounding earth formations and each liquid may be provided with its own unique biological tag. As such, the primers  171  may comprise more than one type of primer, where each type of primer may be selected to provide for amplification of any of the different biological tags being used as tracers in the reservoir. In alternative embodiments, the primers  171  may be selected so that the primers  171  provide for joining with all of the different biological tags used. In this way, the same primers, the primers  171 , may provide for the amplification of all of the different biological tags. 
         [0052]    In an embodiment of the present invention, the real-time PCR reaction mixture  151  may contain probes  173 . In some embodiments of the present invention, the probes  173  may not be separate elements, but, rather, may be incorporated into and/or associated with the primers  171  and/or other elements/compounds in the real-time PCR reaction mixture  151 . The probes may be configured to provide/create/cause/be the source of a measurable physical effect that changes with the amount of PCR product in the real-time PCR reaction mixture  151 ; where the PCR product, in some embodiments of the present invention, is the DNA product produced during the amplification/copying of the biological tags and may include the original biological tags and copies of the biological tags produced during the PCR process. In other embodiments of the present invention, the probes  173  may be responsive to incomplete copies of the biological tags that may comprise the single tracer DNA fragment strands  172 —the single tracer DNA fragment strands  172  resulting from denaturing of the biological tags or in subsequent cycles resulting from the denaturing of the copies of the biological tags—coupled with primers and/or nucleotide bases. As such, in embodiments of the present invention the probes  173  are selected to produce a measurable physical effect during the amplification of the biological tags and the measurable physical effect provided is chosen so that it varies in accordance with the amount of the PCR product present in the real-time PCR mixture  151 . In aspects in which multiple biological tags are introduced into the hydrocarbon reservoir and the primers  171  are selected to provide for amplification of all of the different biological tags, the probes  173  may be selected so that the amount of amplicon produced from the amplification of the different biological tags may be analyzed, determined and/or measured. 
         [0053]    In step  180 , the real-time PCR reaction mixture  151  may be heated up to a temperature in the range of about 70 to 80 degrees Celsius. At this temperature, DNA polymerase enzyme  181  may act to add bases to the primers  171  so as to build up complementary strands of DNA that are identical to the single tracer DNA fragment strands  172  to produce molecules equivalent to the biological tags. In this way, the biological tags, if present in the sample, may be amplified/copied. 
         [0054]    The probes  173  are configured to react with the primers  171  and/or the single tracer DNA fragment strands  172  as the primers  171  and the single tracer DNA fragment strands  172  interact to produce copies of the biological tags, if present. This reaction of the probes  173  with the primers  171  and/or the single tracer DNA fragment strands  172  may occur in step  170  (not shown) and/or step  180 . Merely by way of example, a probe such as SYBR® Green fluoresces in the presence of double stranded DNA so as to produce a measurable physical effect, fluorescence, in the presence of the biological tags and copies of the biological tags. Because SYBR® Green may produces fluorescence in the presence of any double stranded DNA, it may not be used as a selective probe. A quantiprobe on the other hand may be sequence specific and may produce fluorescence during the annealing process of step  170 . 
         [0055]    Steps  110 ,  120  and  130  may be repeated to provide for further amplification/copying of the selected DNA. By repeating the steps around thirty times or the like, it may be possible to produce in the order of 1 billion copies of the original DNA selected for amplification/copying. However, such processing may take in the order of several hours, and is qualitative not quantitative in that it only establishes the presence of a certain DNA in the PCR reaction mixture  101 , not how much of the certain DNA was present. 
         [0056]    One type of probe that may be used in an embodiment of the present invention is SYBR® Green. SYBR® Green binds to all double-stranded DNA molecules contained in the the real-time PCR reaction mixture  151  and emits a fluorescent signal of a defined wavelength on binding. The excitation and emission maxima of SYBR® Green I are at 494 nm and 521 nm, respectively. Signal intensity of the fluorescence increases with increasing cycle number due to the accumulation of PCR product. Use of fluorescent dyes may enable analysis of many different targets without having to synthesize target-specific labeled probes. However, nonspecific PCR products and primer-dimers will also contribute to the fluorescent signal. Therefore, high PCR specificity may be required when using SYBR® Green I. 
         [0057]    Another type of probe are known as QuantiProbes, which are sequence-specific fluorescently labeled probes with a fluorophore at the 3′ end, and a nonfluorescent quencher and minor groove binder at the 5′ end. QuantiProbes form a random structure in solution, which facilitates provides for quenching of the fluorescent signal associated with the probes. However, when the QuantiProbe hybridizes to its target sequence during the real-time PCR annealing step, step  180 , the fluorophore and quencher separate and a fluorescent signal is generated. The fluorescent signal is directly proportional to the amount of PCR product present in the reaction at a given time point, enabling sensitive and accurate quantification of target sequences. 
         [0058]    TaqMan® probes, are sequence-specific oligonucleotide probes carrying a fluorophore and a quencher dye. The fluorophore is attached at the 5′ end of the probe and the quencher dye is located at the 3′ end. During the combined annealing/extension phase of the real-time PCR, steps  170  and  180 , the probe is cleaved by the 5′→3′ exonuclease activity of Taq DNA polymerase, separating the fluorophore and the quencher dyes. This results in detectable fluorescence that is proportional to the amount of accumulated PCR product. 
         [0059]    Real-time PCR may also be provided for by using fluorescence resonance energy transfer (FRET) probes the real-time PCR reaction mixture  151 , such as LightCycler® hybridization probes. Such probes use two labeled oligonucleotide probes that bind to the PCR product in a head-to-tail fashion. When the two probes bind, their fluorophores come into close proximity, allowing energy transfer from a donor to an acceptor fluorophore. Therefore, fluorescence is detected during the annealing phase of the real-time PCR process and is proportional to the amount of PCR product. 
         [0060]    Molecular Beacons are dual-labeled probes with a fluorophore attached at the 5′ end and a quencher dye attached at the 3′ end. The probes are designed so that the ends are complementary. When the probe is in solution, the two ends of the probe hybridize and form a stem-loop structure with the fluorophore and quencher in close proximity to efficiently quench the fluorescent signal. When the probe binds to the target DNA sequence, however, the stem opens and the fluorophore and the quencher separate. This separation  1  in the annealing step causes the generation of a fluorescent signal that is proportional to the amount of PCR product present. 
         [0061]    Using the probes, real-time PCR may provide for determining the actual amount of PCR product present at a given cycle in the real-time PCR process, where the amount of PCR product may be indicated by the intensity of fluorescence. The fluorescence generated by SYBR® Green I or fluorescently labeled probes may be indicative of the amount of PCR product, amplicon, in the reaction, including during the exponential phase (log-linear phase) in which the PCR product is being rapidly amplified. Outside of this exponential phase (log-linear phase), the amount of amplicon present may be a constant. By selecting the threshold within the exponential phase (log-linear phase) for all samples, it is possible to calculate the actual amount of initial starting molecules since the fluorescence intensity is directly proportional to the amount of PCR product in the exponential phase (log-linear phase) and the amount of fluorescence released during the amplification cycle is proportional to the amount of product generated in each cycle. 
         [0062]    In embodiments of the present invention where multiple biological tags are used, the probes  173  may comprise multiple different types of probes, where each probe may be configured to produce a measurable physical property in the presence of only the PCR product associated with one type of biological tag and where the measurable physical property produced is physically distinct from that produced by a probe associated with a PCR product associated with a different biological tag. As such, physically distinct measurable physical properties are produced by each probe when in the presence of PCR product produced during amplification of the different biological tags. 
         [0063]    As noted above, the probes are configured to provide/create/cause/be the source of a measurable physical effect that changes with the amount of PCR product. In embodiments where the measurable physical effect is fluorescence, this fluorescence may be measured during each cycle of the real-time PCR process. In this way, the dynamic range of the reaction may be greatly increased, where the dynamic range of an assay determines how much the concentration of the initial amount of biological tags in a sample may vary and still be quantified. A wide dynamic range means that a wide range of ratios of target biological tags and normaliser/reference tags can be assayed with equal sensitivity and specificity. It follows that the broader the dynamic range, the more accurate the quantitation. Furthermore, because of the specificity and complementary nature of the primers and probes, the presence or not, of several different tags can be identified from a single sample; where each probe is designed to contain a fluorphore that emits light of a different wavelength, corresponding to separate detection channels in the detection instrument. 
         [0064]      FIG. 3  is a flow type schematic illustrating a method for characterizing a hydrocarbon reservoir and/or determining flow characteristics of a liquid associated with the hydrocarbon reservoir, in accordance with an embodiment of the present invention. In step  210 , a biological tag is added to a fluid associated with a hydrocarbon reservoir. Such a step may comprise adding the biological tag to a fluid being injected into the hydrocarbon reservoir, adding the biological tag to a fluid present in the hydrocarbon reservoir and/or the like. The fluid being added to and/or injected into the hydrocarbon formation may be a treatment fluid—i.e., a fluid configured to provide for such things as stimulation, isolation or control of reservoir gas or water—a fluid for use in a hydraulic fracturing process—such as a pre-pad fluid, a pad fluid, a fracturing fluid, a tailing fluid or the like—and/or the like. These tags may be used to monitor wellbore fluids such as fracturing fluids, injection water, drilling fluids and other water-based treatment fluids and may also be used to determine the results of adding such fluids to the hydrocarbon reservoir or an earth formation associated with the reservoir. 
         [0065]    Alternatively or in combination, the biological tag may be added to a fluid already in or flowing through the hydrocarbon reservoir to determine flow properties for the fluid, determine flow changes during extraction of hydrocarbons from the reservoir and/or the like. In certain aspects, different biological tags may be added to the fluids at different locations. 
         [0066]    In step  220 , a fluid sample is collected. The fluid sample may be collected from the hydrocarbon reservoir, a wellbore penetrating the hydrocarbon reservoir and/or from a formation outside of the hydrocarbon reservoir. In certain aspects, a tool may be positioned in the wellbore to collect the fluid sample. In other aspects, a probe may be extended into a formation or the hydrocarbon reservoir to collect the sample. In yet other aspects, the sample may be collected at an earth surface or from an earth formation associated with the hydrocarbon reservoir. In one embodiment of the present invention, multiple samples may be collected from one or more locations—either geographical or temporal locations—for analysis. 
         [0067]    Once collected, the samples may be centrifuged and the collected material dispersed into buffered distilled water. In step  233 , any DNA fragments in the collected samples that match the biological tags added to the liquid associated with the hydrocarbon reservoir may be amplified by mixing the collected material and buffered distilled water with nucleic acid bases, polymerase enzyme, the predetermined primers and magnesium chloride; wherein the predetermined primers are chosen to provide for DNA fragment specific amplification. By adding probes, as explained in more detail above, that are responsive to the amplification of the DNA fragments, a measurable physical property may be caused that changes in relation to the amount of the amplified DNA fragments in the mixture. As noted previously, a plurality of probes may be provided in the mixture where each probe generates a distinct measurable physical property in response to the amplification of a specific DNA fragment. Merely by way of example, where the measurable physical property is fluorescence, one type of probe may give rise fluorescence of a particular wavelength in response to amplification/presence of a first DNA fragment and a second type of probe may give rise to fluorescence of a different wavelength in response to amplification presence of a second DNA fragment. 
         [0068]    In step  236 , the measurable physical property may be measured. Merely by way of example, where the measurable physical property is fluorescence measurements may be performed using a laser and a charge-coupled device (CCD) optics system. In such a configuration, output from the laser may be passed through the real-time PCR mixture. An optical fiber may be coupled with the laser to provide for delivery of the laser output through the real-time PCR mixture via a lens. The laser light passing through the real-time PCR mixture may excite the fluorochrome in the PCR solution and the resulting emissions from the real-time PCR mixture may be detected by a CCD and analyzed. Image processing methods, such as software algorithms or the like, may then be used to identify the wavelength being detected and/or an intensity value for the wavelength. 
         [0069]    In step  239 , the measurements of the measurable physical property may be processed by a processor, software and/or the like to determine whether one or more of the biological tags is present in the sample and/or a quantitative amount of one or more of the biological tags in the sample. The output of the processor may then be further processed in step  243  and may be combined with measurements from samples taken from different locations or the same location at different time to provide for characterization of the hydrocarbon reservoir, determination of the flow characteristics of one or more fluids associated with the hydrocarbon reservoir, determinations regarding interactions of injected fluids with the hydrocarbon reservoir or surrounding earth formations and/or the like. 
         [0070]    In a fracturing procedure carried out on the hydrocarbon reservoir a series of fluids may be injected into the hydrocarbon reservoir to achieve the fracturing effect. In an embodiment of the present invention, in such a procedure, a different DNA tag may be provided in each of the different fluids during the fracturing procedure. After the procedure is completed and production of hydrocarbons from the hydrocarbon reservoir is resumed, the fracturing fluids may be returned to the surface. In an embodiment of the present invention, samples of these fluids may be taken and submitted to real-time PCR, as described above. As such, the samples may be centrifuged and the material collected dispersed into buffered distilled water. To this mix may be added nucleic acid bases, polymerase enzyme, PCT primers for one or more of the different DNA tags, probes and magnesium chloride. The mixture may then be placed into a real-time PCR system, such as the Cepheid Smartcycler, and the amplification and detection process is performed. The data generated may provide for an indication as to which tags are in which returned fluids, the amount of tag present and how much mixing of the fluids has occurred. From this information, determinations may be made about the results of the fracturing process and/or whether clean up etc. may be required. 
         [0071]    While the principles of the invention have been described above in connection with specific apparatuses and methods, it is to be clearly understood that this description is made only by way of example and not as limitation on the scope of the invention. Moreover, except where clearly inappropriate or otherwise expressly noted, it should be assumed that the features, devices and/or components of different embodiments can be substituted and/or combined. Thus, the above description should not be taken as limiting the scope of the invention, which is defined by the appended claims.